Skip to main navigation Skip to main content

Korean. J. Breed. Sci. : Korean Journal of Breeding Science

OPEN ACCESS
Search
Advanced Search
ABOUT
BROWSE ARTICLES
EDITORIAL POLICIES
FOR CONTRIBUTORS

Articles

Page Path

Article

유전체재해석(resequencing)에 의한 가야벼의 벼멸구 저항성 유전자 탐색 및 선발마커 개발

지현소1,*, 안억근2, 서보윤1, 강현주1, 이상복2, 현웅조2, 최인찬1, 김경환1, 김송림1, 이승범1, 서석철1, 이강섭1

Detection of Genes Conferring Resistance to the Brown Planthopper (BPH) in Gayabyeo Through Genome Resequencing and Development of Their Selection Markers

Hyeonso Ji1,*, Eokkeun Ahn2, Bo Yoon Seo1, Hyun-Ju Kang1, Sang Bok Lee2, Ung-Jo Hyun2, Inchan Choi1, Kyung-Hwan Kim1, Song Lim Kim1, Seung-Bum Lee1, Seok-Chul Suh1, Gang-Seob Lee1
Korean Journal of Breeding Science 2018;50(2):104-115.
Published online: June 1, 2018

농촌진흥청 국립농업과학원

농촌진흥청 국립식량과학원

National Institute of Agricultural Sciences, Rural Development Administration (RDA), Jeonju 54874, Republic of KoreaJFIFddDuckydqhttp://ns.adobe.com/xap/1.0/ Adobed     ! 1AQa"q 2#w8B36v7XRr$9bCt%u&Ws'(xy4T5fH  !1AQaq"2B Rbr#u67Ѳ3sTt5v8Sc$4ĂCÔ%UӅFV ?_Aנj- H>>,m*>fzp"TrKkr^r.|_&]|*vPuܶvoQ1mwVJUhu-I"=LniAƕ8"۲ k*ҿ[yu:.vUQ+)%F DHyVBk>Hy8jݹ q~9D4KRmzQ)^ʔ.J%k_tVi5NTjg!'ky|5asOȻ)R۸ߩFMԿ3L4j6dڜ#NIwUF]JqB/(FafJRzq3\G՛ ?~\ 6)6W4m[O^L0E&rRMض*C .]Unl-1 1r#Rj/&QɈ׉˩s6Rj=5Tg.y.·Pӡ:JJS:C8-2u]d&vUz;7p9 5VnL֢"y)">iי(IDDd| Yj0; LRfS:ktYK%*N2^m|&dğth":ey)uPQZW)gcC3Pv&MMWd&Ŵ۲mvTRoժM03*F3Yd6\8,\hݻ kߔi<k NTwSԪmljj[>->ptU%'LR>&EBH$MQAUx[$Z6vi&_a.KIQ{hyƒ j"JOC9eFҝfj;˚Ω<[3_m% lQ@4g=5$(J]Yc-OMq<Ǎ wSzڗ)k$7VIP붾ͯnV+卵*t]iЎD31~SA1éC2u)ʼnQn-Uoi3:grI8ؓWm*G zܕ)ZקJ}Y YlGeJ6cB2I NS3Q>k=KTBT]W6+SOXQgGR? telˊ%-Re\hѯ2TF"C/OJΩ6r[N.0{SpljjX1“jOsӥ;ҭhe}xu`Ք&.)yO̒ Fߑ.$Qw;9Iw2o+RVJMSOj[SoҌZ%;`d$blQ{Ro{Imڌ>3egf\O֝Uzx"䢸g+mv%Gʆ:|V[N'&ס-ޝ'kfE|K,G&˳98Juin/\\Qݿ̋v~Ǩ!rtWU d|E߫R4d}.qPw*Ӭv5YEcn~f5c%MTMkb-F>5JT,})QHg%{("ӔȸWMsYyWNRrkkJr0XドnͫT}r-jj,Ŕʍ\Q2Ri>v$5!]"JB2WɅ)]VԜUc8i|.jeRO6^V.¸ Q&#|ܶ-*uOG%JAtRZRr]FFG\۩w+?'zչSѧt jz>KW&ot{7P&2D;&\\>Q2JzܗAKSfeNn[jRrԕf6,q,F1tRfԗ>vֶևj-&R'Zi2=xv~Elbsvm8=ӛ"ū񕜈BȩlWau[]ٷBߨF~J!|Ipr3R̴#Yp)={7:G{+:\W}n|Q#%)7^-h"Ƒq:M*%J&$T軨I333׎g_- ucBwwjp[6i25$̏bU’ٱRv?G\~#Iͪb7<<}Ezt" q_Inw,7-d,G÷%T* Wg1"䥱kq/A.,_KhqŒxwvo u2ۥۧ.bQ}XκA$֣ +K״ZUNmڸII{.v{5z5ѮRme[moyƾd~cRݾK'j.\i&/S6f|b=5: p!6i_ 4j6=.si˧eƾtS^c.Y^RJVS-Vi3,esi08?H$GvZgg?gi䤟2adw릿:"۪lkSN>q-4kI܋ێe̊qۅgDoѨ9; #T.Q;7#~_Ufstb_'w~Xw1Xk,vcOt._}v}8"(4Z\ۘgk?J?bm_c!g{HZV]Fkk%~gEt)b秴vΰB|꽸}mp~E6ݹv;7P٤v+ri*3Ԣ|'O14_~7nP{7ZU\Vű[ +7󖱅o#:ǥŬ\|3r%TJX]V7ez¨Y]lc|O3V! R zbJ'PnGqVJ"19WVeOF埜EaEJωqCN5Z g-9[S<$sUK5b|7sn\7x qmv##FF\ w[=-43$^ooVSiXօv7iB۴yg>]Vf"r$J3""32!Zh[K%7GvNLs+4nB/B{vlsobJaҺJR:0g%&zR\ S3T[&ִor*ⷳc3ʊO[iozW٨%$gn:ܶWwFBԹjHP&z u&F2\f;ipW73 [; '_̽b;vib!oec dC-tS__$Xs]l9&z$2/N>%'[}b{h/{`{Ji׉׏ YJB/X%}.|+{(S:qz]4_Kѵo`^tY_4S#* ^zvݾMr+TrkQ g.8Ͽ^i>ӈǙvix>$o( ^qt*&t1oJVu-ql5U6jCЉmĻ*"?JT=K'O/|=Vo}l0b}}f?X[?/\JSBe,kP8ETJ==?.p5ފgbU9}ǶdNKk—_$8̸͓ۍ8Di\BԿ-1v{FF]|.^ۅ{vl12׏z7-R7wE?\nh\jN/Kձr_oBw"N QMBZqe-m:ӨSn6j4%!hQ;sv'm4kcM=!8\m[M4{SMliۇ%eֽR&N:{2A8)THLK3Zj[jPBx#BگMf:G1\`edcʮ?|w(-̮vXt,bW2;.ιNHRR#YwTM"<;mk\.foIDjmlJ;vxy7o7i\,KQŊ9d^Mmgc L*.T6tLeIuOH3SJQ3=F/ʿ<9\JM6mN6=<{xkP!F1QR[I$6ُimXu2An2yԒMU q f[IB-'䤯jYm52&JG\zд\~vdg QtHGXw&1Lw+nDEdC1w|YJmvP)HZ>i0BPβә?R:QO["]I_Jʏۍ>QKyu^bycBq4lXF~l [\*N>-J6,Gq(Zr5h]CwYӤU~ʶߑ u*SIv%ZfJ7)! FS*s_\|IŸZ)J ]ܜi4"z[+Z,MOZ))}|Ʀ(RUNIII.S'ˍO~˨rn}M)xxӕ0 eyҵ7YMAB]ӣU:/ѭ*6bcwP͵ "+qēVjŹO|GtY4V j[mLV M -m>",B$ GD1~j6O4|LxnNmqATNR3ε|DŽa[fmn-ڭ+FiK7Pcm;r5 l8r{#-]'nrFh2ruycb;pW=njRqRJ(d mnpckNnʹ+6]tz~E=ʕ l ZZ5jSi3#47.Lcfe`9؏v囜.F\-UZ:*0_<Νu9Lӵm&)_3\^ҹ3"1n1v_|uRʞͫr'iȧN_kH׺8xXrj=\МH)V\ˬ.Xʸ oVRC}ySU9/OBY먌5 ٿwޞ)rw8Ӫi5*5ZΗcGƱ !ZۄlmpjJ -l <R̵/JAպZuq\IdUS 48wXJJtcg4cI~aqߓwŷrm-v)G7yS^7H^-\mŌAq|"m9IBnF㏉9[N+mmy/!KKۉ%n +BdddfFF6FQRN-U5;Sv'm4kcM=Mn)\qιqUd9F%",6MGdT%-+~ f%+y֛^3SrF>6lc(֪vۊN;g._0Sѧ]ETWرkQKzGe9ʨsKA"yC y2\[5 rԭ7Gk5Mzw_4sM3hxЊ'oÍ5jsub )ͪ~tR2H]R͍>̋m6=%(˿(Wrr-܅y5(ܔJ޺YunW̹븹NsqK ]/QR#"ZMDfD|43Qw|._ԡSqTZBg??O Ϥ)/E_U|i}2 9Z?¹0:x'3,whǣ?C y-A~=daJј&M?D1_PS+Oi&;a @;Dž7[ zZC"bv:jjMQk$M RԸ3uA\=wI.AwC"^.{?-\NSiˏ"b}T/}q/ o.1M}R%:-ZniʒL$SgrBW*,Mw'N\ɇ{s\j]VryG'8f`}'N<*/`U숻z CwHq18J+vԕKss4R53/&XTt1bZƟo\=%nO)h$rBi-nKĪ^ ջڜlwkYm[̑+/QrZo%TQ;TLs($2C:s.%+eoNttq۰kK7O0m_t_pZ1SsSM7"mevFZ[w -FJ*T*jФQRg BSu|]g:ɵzjqwmltL.e3sRMچkSmjkmWœިm++¦'tILk*բQ D,PB\lI[9{%Gb R6öۍmX-MaʉA931cs..G4CujQտ[9 }G-xwl)IQz j Ó"rqe&=]꾧֎c)<kӳ+0JrRR3'TnXi^xMF Bު*tIL.[h"2"nKzZe'ZV/RrNYz]8죝n]Ķܩ>^Ժ]u-7^\mZjܣ9+Rmn ߑv?oꋘ?&ƪy^N4o=3-ؔ̿*`}V݁ ƒPu8%$ ݗ]wt;\y\>='OjPIp/nJU8{϶FNMsf"ίNqƹ(+ ݮF2Km |jܴZs%zf*eȫ?]4)I۵nR&FX + [jDh(#哑9q9Eծj8noǕZf\J-l&Z˫}`ӎhyrΉn\űn]9pʌӣ"׮Wt?N4_I_~54#/my1Xr*척aS#DT >q ssΛW;3oUaJSRMDgQnt:Ql,/ ܷfRqiM Ȼ>Cob;A>ڦWقM9X~/!'MW.}Vrߔꔵ!5|iB(0-zF=}okڢE$^wW~nokY߮\6՜̌{i-AF*9)\t9IV6۸5ZUF6R$ŨQIq砳YUZ]eyv >hI櫥N )&l JulwE1GDOuFN2| }馥uC1rޫV+^gdb&W[4<^e4YW,d|htͮsUM)۸8:{3d{AѢ)~ \#J=NdƮꮓ90 |1K$v*?мS ]i$J,C,SG?/_՜pMSƯM|mG1V1$~K>CSvkuj=&) -,yLjuFHK{c駗.SOua;BrSqj-ۍZ#'Jys7[g2z/.u4+XV2VQ.ޕ)$"(%)#Z7suZ%j }BǬݕe)Jvz8zJf:hIN|svO1O#IEcۍjݽ:SdὮvu^@:o^5cs>i/VqmVm]ؔܢn6'vޑ̗J4Wn@OlKbX ;n:hgJ9ŻyǑz8f܌q&Y fN0N;[69 rbׅC2/#kE l&2~èMR.*%g=Ft.%؝e8<.e=Uv{~㻏"EˑnvDѭ͜Lu3u0:U֝$[M5<:oi+V4V9 6nXvx&_ q Qqw3W:uϔ2yb/(ɳ|5zQiJ#r|Hw#.W?4aDŲ\ugWG;Cw鐢K|xg)##=O.dF˟jMUvWĻsr.z]kPc9"]R)mkfOd*uYf١RsB Aîh=k]ʳUrrZsq`d#r$/Ը3o^&lRWȍyuW̦Y4QDUMJ65ƒ[+ygk XK_±k#y:8(TJOSQhJt2.DR}"5[) r)6V6u5k:eXZmv𭤔!푊Q[qQ}ҹLE- 8qIZG|UM4j}Mܕ[Vwm{} Naqµ"ԈM zOpKѰ?IAD3Ir0'/q1itoB5{%wkOBn-ۜduqIzYK60{+DʕܞqIt";r1mG/\/ym[6JƫR \L=S=OT@Ix[TMm{>ݾտ֒ݸӉLYIx>+"JVNzx||5rI?C{oz8۹e\R-^\A2F R+N9 vlT]"ۭ d)t֞i #E2jB@׵=#/N+!ĕhx}I!cM`ąZ*ŻɄҒ߮Y.Z}='/oۙ3IpW̮hT7cTSuz9>B}΄&h!>lӵn~j˅IvU.'v'CSZw8QK3G> ,J59ٷ+HSg䧎hJdzvwv-cvxS5[̊n~ؿ%ַX?O0\6ne 6kn9.ϯ} *h 8_QhLݣ7q +=XBҲ5?[[)+F`=4 }B,sNg==u*Nj9k_GJ)+R~GSPBȒZ:(K]heL=vKPӢwq(NrG^ثϣ?#tC?.ͼ[ۅo؞y#%ǛjVyLSw%T*s92JTM%"YkQО.q)gCͲn8cgi6j1MѾ[{9h^vƘǚםidfi.^RHmg&rׇz:}݃}xT$ضk'5s-狶,\vpbPD،=Okf.c#cdz2FK5T!&)|ntD<+OŹU i-G[EE*FDfeaf2QƤM\UG_{ǹm%\yrGy:.\4wjPGUJޕUV7Do\7Vy_13w;[?c]H\$IJ,*L]3b%L{y.JRKG2sq,B6T}(#nW|km+q5] r㪍bJ@y{byz,b踊3ϻJ,'^xd،)JVw#.Vټc''ÝպWtbRؒJz۠8!o9IۄS95E9ؔ-e9JR{dmnｯ<[~n${~Њ$W?&ՐY_? #a.ߑv?oꋘ?&ơ|y^N4o=3t=~7!/M3>n8W홎2M`Qx+ z qy8%]7_~540ۦ彷]Wq CѡwkďyF5Dum_}~P(5.(X,K9vᯐ?leB9;Jhm#3{CxGE-S{;@Fz˙]=O'!ɿ]' r`:7'2bЖ>Iy,/eTy/V<.H?UYY{\^#ѣr9^7?xoRȆ7EoS_&??zϾM?(~Q-K&>"~aߨ t7Emsϛ+?;fCr)fY+>z$tIkjn_>vnrֳki-˹l= t;'EyC¥|/BLwBJdgjۛ$s S1|ɍV%JI6KvəhzIlBYɒ|0"Sy0F>eo5W)O+X˻u';v)2vVq۳kۮws?UʑBǴYO漪e2MIjPAک\b1)DDؚKm6ZWΨgȕ۶yjڳ 2ضN[C[|r@9Jfo<_eI7q.|cÊV߷:i.:$ȋ)1%%)ADZCEBxJ0MJۥy(bNsKM9k43IwNt.\%N簤I'.j|ƃ2$grBEٌ\}9:v*!n7M(ɽ]7c@XxƱԨ37īf62cTTfFK]9wntQHͮvٱI/f|j=7}\_V5U^+:uljSȃY(XI.ȱmo1甅jڎIZ2>#\*:gY|4k\8ZwSqtyA!+];бޞKծË¥e)#5ap.QK^8VdU{*ѽL\=qmjnB5>{ Ӟ`v±5 ^k&O~Oshɷ,;6nOW>u6{RqS`)S%jp\ipdEBLfTWy$GIYw~䲭J.1vSY5z.V>^+Ǎvc.I[R{QsNR3ӎfhd>y?UJ*}~[e\i5U^͛E]G_FS(Iɿ]i8:4zj~շsW,ˆsy:%O}iur]iF5~3M:Ӟ#N06)4ߧgdawIotiz:1r5YDZLHBSi;NQc44la=Y kQIT*ըl:tq2(է9VO4뒳܂~2rq'nrVZŦ[t7\oլfb/mlpc.I8콚q^1iE~䰳mi[dۧw֤ICfdFeCsg:i| 6擣׋* 96lust^{%99UNRvaMܽo ammi$em4D6DD\nA%$$#}۷/ݕr99JMն[oT޲E"KTaP+HGkŴj5TM5xƱOS-k`ۛkٝWz;{kS}F;~q|~^_|euwnE'pSupUP)V]vE+t =ZRaVdG6= *.ϼnj9:UɷbېmF_tޫgHjVS'śǕًdkkѻ_]Kv?nT>)^e=Ar1'3ԔILyD?:-^in):{7.؂\.:V }#뺾.3r̸*xbFM aȵz 6SQ:ײj[ 8nn iFMw rR"5M5I旘35f^j='j:nNW.ʭocZvZKV^ɚJ.cM1ZI7E'6rg탸5oZ=[m Z`\hbMUR١Ȗĉ):Jin!_7Dй+f̷eKҷvͨBPR(V`y6tw*MRΝcB.ڭTnc;P$8nFvm4(D(R#R-L -2:FP lxZKQc6I("Km%$E, 78uXIFA$RQI$JbInG]c[ֹ:ZM+n^')JmJMJRu{e)7jQDw~%yQl}BZujSSf۩QZ+Dzhd5o%BIc'GZ?}΍:>Ɵivז-%݌J5MqGWTVʦh݇ܟ~Օ_6 n'{3~mϬj'J11OȻn߃r Qr\3y٘+WӍ'WxEs^O3 o~[|7>]]H9݇ZomT@]?5B:Z߂'`V_+/MSKX߆ޠk3?o7y:4R/7þ] iG߬aBRU&?r&/} cQߥGj2?C5Yśe7hU=?+ x龳f-܈czW^7p%-(\D4h{UK&ӡn^m]Fݢ:`δvj俜F+) y[{{ 7 tu>gvrěOj'5 iRg[ͶFjGe n~qT$ci ۚ0oԹc*jL[sVWqj\ݻ&6"WoK:cnWmrv)o>66(F>=W^bf#c zzʞtپy%mՉPël e}J.\Zk4ttt>oEM=q)hJjI=ͥ(%]脼_88ф;͛gWG;Cw~˘$4=uWdĜTثNDkiQL9U*O"4XP`02,Ge-k5$h>ܼ]3vr6!9RQPIVSnM(ۓ{>;/Qͱv{3&-[rc)ܚI$n{Sv3[j00)-D3z}MRzVQпj,T[uVs0\}Sid;r(ݝJ>æʺL&c[jPK0~d(FKÝW\m]GTcF|Iׁ)I3~#oX%vҦEݑؼ5Żv2qAZTE^..M{ʐfȏ2##.R}*KʛZz^ӞN*lPťLf\G6[WVQquV]XAi)5J!,$iJ6o$tPZc;Kjx_n3`qIelV~vLy{fn匋Ѿn%;zV.n'-ұdd2߽1bZksPe3TI9)$ԩIN9Vơ\=2885N\ p)/a柛w9g_lױo8ݷ iixJV& ғRi{N^_oAŮE6Y7I$Nk$|Q)-*4Z)^¸%4Qm [I%.c-OV+C֧R#%ѨCe3i;w$G+_dy| Fzj$DI(=OA gj%v/]8qԯNIS*֩',Q%\44ZZ%D|Ǧʴ6&vֵI$%8(ԬƾS&#Z. }6z?b/|Jl{ץv&mpx4Z$”ڝ4-H%dGKfM:sKSRWeJAn]>s6应-W9'H]'uȫYvgK^\czp|My\鏩w/ËQ.)]\QiS`8uL뚛̸=J"ܻi\å'-)54Ue]:K\퓡vK xwBqrH\*֕TnzC.mT=t-H]SČ~Nu╏NÅ3f|͡G~B+Xm[Q7U{9"~jgK Zoʰ7"qJ,ekSeNGgϳ] ^.6:s}_,%eRg<5⿨z{ZPun#jRІ.6g T.!]xa c#jN$Zpl̋H WZu8WmMRýsĮ?Mco~sx TU҆Q :KDG4n42.<3/'^?6/ܠڒ^yrrÿr2\D}}B]^E~^T cɛ7϶Y[<֞[7d}2%QPqOLEQR\CIsj1?\}%tJ0e~ *sk"*)&ۓEi#{1J8Hrt|'ܝRr8)=ƔN'RVz:cf]F7bZyZUȘ4x8,#JG̒?.W9XnO]KO]%]ƻ O5Γ/3qÓj؍/r̺rƵ 5\&m6h.xoeX[=<3%< lZ"2h\Z[&jW3ejm?k&[]ųj+{N{66leu_+lj]q* 7g*knأYv= q ەdxЬZ|%GUrQ3jLŒqET]1% qkXYūYc[7Ś]QY\jko\</Lc7+'hMSUc6qXyؙ~6#ѯv.0$BQi5YyIhɍiy=KD!n3Vm[V%W-B%swa97ajۗ m+9~]fKq|Ddaˑ0A]_v޺mM5* F-BYHJ5}q>ʉ.6hyDmpD׬'-_v5;5[8K[viJ.3dR:oYHHh9I7:۽fi+wm^ [)odPѱ52CZUJicSw\&_s0uBȍh32džzQflcd^m|7GѹE!fO5]]H9݇ZomT@]?5B:Z߂'`V_+/MSKX߆ޠk3?o7y:4R/7þ] iG߬aBRU&?r&/} cQߥGj2?C5Yśe7hU=?+ x龳f-܈czW^7p%5|Y:SJE\U-(a_cƣUǽXXKiȞNlmۊڭڄR!**ܤMeȽ$|X5(Ź\rJ~ܮ]>'HB0cp XFr_c?f?7<ukSgov¥iG>>䙗i.+t+bOjIܶ . i^:nm}s}(3>NZ$2Qg([".>i.ƾ)B̋M8+"- >eE6DݥJnJˣt׻ 5.˅nJGwZD~!i۶a,Db3ZQ3O#KO5/֍ozuK'GbRi᝘NV_ҝcvם ZoX}F6z 7e5_e:ۓj=AB+iܔERadMBq*ԯ DwI/Gy*mĥiRKg6skY/#SN4e$-yXM YL?^ĸNNӪ{$r1JJRSLO]Aqm>V/s[~i/j+m>z}eI"Qvp]{ZԼ:{vPAG2=T͡@ڐ#u"E*>C;o$~C#_d/HBq^YRٽzIKbOm\~żjFFGdiQ(*/i*#.FF]©m=BmpQQQSP&Ҫ!T&^>:y)$ˑÐFčI Bӡ-t!bM WҦŶ'UZ=}zvn~oT/\ǒ'nr8 AJIӆz<^uߖ4eFC1i+v!3qNyߕni?4JZlmYFXFۼO0B\m[ tʄU3s"Sr(NJ;SKW72L4̏BVdf^Ҹj\]ȱ۪(ӷm?J-KEmWڽ^4<8qu%9pŹW~877ܾeVгS(յe^C]yX͹! םm4FGȋ\y'Z FX7e)|Gjt߹#gb\ŧq_([R8[qU$Z (ʻezV2V!iQ,i$JE˂٩ a(GK'O{vnBvryRd-RK4=qxZJMl_CuuIz @Rt㮽޳!|68\-l[џ84-2Pu" RJ_^OL>G1~XnBŬw6J0*Uvlږ1N G1q9IUm*'oWu][&UyYZbBZRZNfEJf"+2nF~Eû7n1xv.RUM$6 lAxSQJ&n5ܞwlEói"#>4׿Q.nEq7Oko[1wg8ZQwZYiqtm&~">Bo?w͡ni2峋NCEy Ҕ+%ZJ ʩq*fpˤl,~^Mχk1+:ݕ z&Y`KLӪУDr3[*Z :(SL&ݻ۬Vqsyԭs x|iI߽zZrg.:mp%6ԜvgmpIUt;QbS.Է) ǨKSV,*lڌ|5Jt3#NP.=+OZ~/G سIgbꥹJnl_DUM\iM!֔wVZuԺ,yV.Q>f v:݇WiaŸN5Ҕ[M7SsrvǣrMW= \8ZW-jsnڕ.ZnF2qt ً[ٻޘY۷Zm"Jxr&NAfA-݌to9s359݆mZ+N1-qS$D=17 x׵+%_ ve4ir6Z$FDڗnFtOr'7'{9C˨ꤡaYoace{Refnft RR"4%ʌm:Sj3)OdInTO>X'vxV#jܮw9Fog;5.~Y5\~18YQܹvj4+~t7S ﬕs %^۵ڴDZV69R^Y+rj$ԇoJKR5wB9C>Y:l+EǎS{ʲ{T6Wi* ^^9k/y/Cs\g*qڵgn4T8mERr|Ti+iPe;;.i\EBEJ 丬i9ɧM-ԼsGDrZ>r#R>~X9y4b棇9JwV۔%m(b[Tjvl}۩~nDԺ{Zo-YuK1vx.nWuO+jN [ٮ0%"΢CdTJK-RަH"$I(*ve &҉FzB,_Vpqp9m8werv')E;o&QE׵^d9˦j\_,ڵugZȻ̧8k+jK{wmr@3ӭ2 wFkzFVqs1؛.v'I%$[iT]D5Dl2 nk7qUxԫLS+sا3/ΖeZYK<["%-g/kRs:f3;*E ت wJ%)5&+&rw*霣i|sMҴ|;R+fm䡩.!**dӶ-6s6,]zAXMWjmnz%SJߴm2UXw7MQ%<!tKys#P,W>s;3IYwx<+i_\\\U6 u7P|xbn_k&ӓVOe䦒 VUr,-㘘"-LZeOSҠթrEvq8Kf%5%&K"#%vD/.ZYYŏ+p$nZkvއuW9㓱Z G wYIFyf)?ƎUm5ԉ/'k84{KO:rQI}XRuԪ|*lu)3qZ[mSm5R3".Xcَ5c®ࢫI*۳~wRϿQWޝ(EJrri&ۥ^ʶ齲Im|[yb;mnm֩uiܘq>E+Ikx߄3r33-5𹻖09ϖ9[Tz~mr5NsWl$oPusޛ^{Z;);sڹf\3oٹZmԉ/'k84{NO:rQIBø8Bݱ3n֤DiK4u& ofSȒܩx<˘|N0Fչ]qsp"}! QWw@t4ӭ+cO5%]'*{eM߲DRO1y*q8w++e!c߶ܪlZWّM欼 CQ̼빶lX{vib/V/ ai;x6~]+z]MWB>re-:lgk}պ!#9?%܋V-c[z!W?c7YNm/jRr[HOzԻefճ0q15Zp#rkQQ0tU-AmڵP/cȕ?0cZYj;:0ZM=D6g ?'UN+ձ[K ܖB2'xq9{|۫N0ku 7xaj;n\ 2[VznMlWiKbSk))f..)Km)&bGZ=>OR܍W:j'rM'wYz&/鶧{Sʵb"vջq[I-ՌZH._x*BagC'T(Q:$ͳQcMCKy?3g'ߝqnT);qs #ؤZ}OOI:cfnc8W~qy.;^pVl]Hԓ>^H^@7-AA܃nmL(uWܻS߿ Td95Bdh4t6*dDh!EhI[iŨ\L.&Nc ܮf^;$R)\rip9I|ٺ?#R.ZDZ;/]nݻqs\QE9M&Bd ]N mN*D>tgbK>+ˏ.!23]BȔR1ɝ^j'k2ƮqBQq[$di]icV/e`޵B.FIIJqbi>Ӥ|p; 6${)RU>_e}^dzdfzi %ekRVUS?6'hׂ)5.\+qUgzE2C˷ecŏ^֔ibk shesFWJ#~> Wk~ݨ}ڶ>ơǚ)׽ZƉo~B-ڼrvoE:Ʃ3ۣK7+Y`WirS):{>ڛ}:wԨ(J_";6R%[u&ƫdZ_\'np| RJwNeTW,=rrbnkڄ[M3ܴz)3- R.?:okۼ0TU'w{6&w7j1z3ON'fGoO?)S_bQ_¿R(^ԴԴG.EtMڇ&RUiW uQjU> Kiu1d<ѥIQ'RQ1:O/lŗᏩiʂv&Jc{D5 Tt)1.n[n۶X}RjqnOʽ(~[Ns{ސ⛌uO,kgo֢dRNQȄ .'6W!׌P朼tdZjFGE"]K@'i۪N;sI[{SOzk>`rRR+!σj8&TjlvA̷Q?HyjyLHNտJMjܶT۽lG?SnKN%<‘ nq[N0Sq[Ta(&t(|HGO~gvkݻTR4&Z$#ViOY1r$6YF?e4U/Mvxų:zbU^gQQ+NW_'4jfz^c'#`rvrڡ(IJ/J ݦ6 ]-CW |_{v*_q3^DZ}Ic6Uڌ8p7{crZq5ki`)mU6|-Z5^iEz3P=:Cu7DF'k%}<C-޹ֲ̱#\,(f88%X-N(ck0VLR~} G"-8ӏ/ϰKq?(#nrVTmZ;zióM4 m |UT'C^_1X.gXM{%ʤd 4\ovN":"y-,T)fLQgۢr=/CƹǨJVr[a+!rT|%Y\ٱzsS>jͱ.oOc6f$q% ǒGo;n[];ߎjrk{~\VۓNIGn:iqxo |~t5)Rxעri{Vi&NUOl_ѮMfsޕkЄay.0P{7N((BaIP$ K"U6Gl ݙqJRu+qN$ m#*p<|{:>-Ev=86N*MM긭U*uѾ?/^o7;'u,h4݌xښRM:5.(/ \իU.{F^rmF-Jɷ.>Q"[4xT^OZ~mK}T0ݛ^SAo9u?lX(' qj%=X}"^e4wˠ|rܫ 6I\Ķ;Ӻw!'ڍWg{ i U_9Avhۣƾ+:vs/MK[ɭīe{`Zgb}r[i'GE2J7Nez579wRq+Un ]J.cJ4M:h箽Wxxm^ pc\wcN%'My $$| :$Fqɏ¾^қP9J6Wxvu}ݵP>Z'FFdg"-; [¢cmWkÎT8nG%ݣ7*\խCLRYZͤiD&J#'ehbSyXK|y*ӞpS̍R`[pTr/Eg)K+92{_ n3zwz'oŸۤ+sOj J:`T>Cf*lwd\fYOP"R E֢̔L4ɥ :;.b(B02rJ蠟9>V'9M%)IqnhP<%,r'P/vNSwr#w"ݨaqc(|{kd=^0jTMR2ULNz|.<|^PfY22##!,K~E BEJۜ&jRNsHަޛg\r,v؜.jK3)[EJ2ii{KEiHP^&]Gn8x=K}Wx/KI9-ϵwQ%spܾ[^R}S3$qvq8M[ ozKxcqmJ/ӿ{_}7&ݨ\f6ZSyQz& 7ۉ[8~UNn|nkiTB+4RI8'Nc%tn{!]Ȋo.nEmʱn𵵥J A+wy#+ikǒڂ;՛s85'KmE:Ђu""Iģ5p=БbTY-ͽڔ詻ngL2Q}$de# fs^o{DUUsfwӶ;s1T,ǤtޒQ\෼J=.tKU,7čJ5 N$y3kdSMQU~mO[03 $zAڟsF5^뜞"Կ QHmrR"ӳηer+ҔZ]hE-6Jmt'ޒ=O[sQj)6K}?e4v_KfZheޓ=BV[bY}lݒTTЬ{ȫvO_qpRApVŗ 6ju=*BR)g "O1yhb=tqJ gtm\b3RY+JQ^Ō֍\յ\>+uSi{=x ^w;uӘ#ĸzLn*$anok߷CBӷ}5Yqvdž<( "_OWit5:EZj2 B ρ1̊fi[n!HQF82q1牙nqnEpT(2RMoM4ϳOu ':֧_Xjsg jP^(ڙ{2%E͖j^}ZU[Q$'U) <܂%!s"m R'G5M0<+zM6qYm$ڕ$3ǧH]?o2N<8F1̻r_my[Rf59NjpzBnl7*{.QP 3N&^BLJPjAHCK2Q}$#~YMq8 k(MFMU)8MEqTy+Tʞ-ar5yܕOXw!e;q-Jqܶ䓊Y:LC UE{/t>r"lI9)3KJjϤA 6SEE$d߇3KG*En|P\ԭTn6I-ƍKTj<1H_zwGr19wF N8ݝ+a9ɫM6mhePi%mmD! """"""*1bRKrD"vnrM۫mmĽm]ӡiG~e"˩ lhRTMk^MX["Jݱk7_ޕ*DqĒ&flՒ}`W}~SմZ{ĕ~wm*/{{ѹ_-0ط#P]xlڱ~Tn5wi*lڪ (JxioϏbqKYR|!|KN53 OS222$jzww%i}>N)E+rۥ7c$Ofl/LNث\6H9: FY󡈾I)fB֔JI_ ֣^: 9mY{66㒢7Uj]:.-os[R&gMF3˸#໹kmjq^8W"PΦURjʄWa˧T!͋ lW48JB2ko+ /Nw QwQzQ ے%$ޓ7^YL|r7!v%Trܥ &|M8~ybrn[RV gSn{{*#2#ԽᢏӴHak" ӌcwҜw&RJ07ױ>Ļ =^ BɆ)v32.M1=#6%̠tҤnzqMwԣ~s*%-j|_m*.Yx9Sz=)qE4 3pk+,`=kNRڥ=B=nŔNAx)Q$ԩȧ4z3t#Z2lҮYn$S%y- JzGpu|LBV7ZW#;Wwipܷ%(6jFG5#{$D"uۭ~]֫SrD܃fҎӾ+Tu>-ZTQ& N|$沸ii>eRWݳu'[O̻j8JۻEѩ[]vni= ڒ,[_%kC7I3Nv$4ɎЈeٸoUu:[}Do5|zNq=Tre%ɧ6&~DȍF]ƞG5q m]/w/ \ʲr8=oʔe9U(W"|S]uZd#?Se[W"ֿh][-7Nu:T=)R}.;ml*5Dlf $fF(̏T hiIUU4Szɕ t(%_|2 ~6eM;TƗK[f&]LK^CE2[ȏBOd;Mi|cx,^6;sیGpQ\NuJIFTJ~đArh* B"$H쉩eXPRj?sl"ԥ)su]xpԴY%VESH"ЋJǰ K&5^Ukzׄ8kEgS2h&Se\ Yl]WҶp-ZUvi7QS:4byqOo+[̺腋[6-_Fo.6[7$p&^ _GZԸߍkc.qqoI[9m߸YxOZЦ1uoiSH)P9Uʄjcq= S>֙NeR><;+ڌk%_qT].srNO?s[=vH[]RZHRMtᩗVؾ:/~u)ԍdg%=edVrISb{6vSu=(ܥ)mTv/J}̇8 S3ad:^hBSf؉OɔLhI_1d8,L><_A0y3rXq"'(۱;mFNII.v5_(^q~X>y{3צ I*Vܛv/jW' T'NR'j%ꔩ:mJ3SB}΋!-H-RJBТQoedi9tjENenPpke.%4]#{:>mkEɱdYWl\\\'nRM4&U>?Ќˉk÷!𴪛]]5}UqG~ݏI"O~s6(Ļ)qO~h}uԕd}Q~G,oE!&G&/]_H-O=o{k\̭bkv.Ô܈+;arZx)m?M\3lU$mk-CFXjTv6u' g:Vn_*qk:VC A%'4JV%EY)#BғO4<e׿jQQ]yUr4=wm[K1r׵%Iũ-O}|kC;/VcݩWZ)EHdžTru]8hgĵ-;=>U_ InvTm_jBM+QiF"9*{DI/iuo(=TzϖmPQl_v4z>T*ȴ>YF;ε\t]EH4ꌇ[VrLzef 2T^V>g2~kg5~Nק;{~Z~W}&ŒBӿS2$J?~(Yœ"˲ߩ\O]: J׉ښT{mmIѩn3˧)4LdFZ/zUG>U> n 5& ϴ-KJi2o]uKljvK3$bԔҚV旧iY5.ίfi96v7!v))FJM4{jG~Jt/lUE%pTAFe4qQk\ve۽/u/Im+W')v{\-E|Pms7߮DZRr۞/mu*1ՙaB܆ -xg3#6ۥtRogʌU)׎]ZҞNnŞr}F1Nnޞ;cZ{N}ۿMiuxʉ*3qi'9KHQ$WJxXyرŔe~[v5~/jN9Q4o6rJv FrdxM*iRjMzUinHdн7ᾞS=S'7 } ̽zt7K|_g J=Lq+/Bw_\ۧx\HJUPzQ<hqF[V0x==CsU7q|^ {)Iq38$_A(VgcKu06Ƅ"%i~_ˉk QCܣB8Ku/񋇵u([w}$F|8TՠI.E !;RJ^}MɒD_q2];Ɖ{5}*n7nEInO{Mwv}&q+v [V}Ĝ@%>#dXQ$f;iep.GquixVt x6bj͵mlKقQ[T]zs/&yەnM'W}!Fp_d^Tu N{ɻ'l{խ2.sTu{W^H&;1s)Pӛ6>$mě;Łnj= fLT)>׸+qReɴ[UR\L*P/!$Ӊ3Q 'K=m~6XqW3^W+ųO_[F$rR*u"T%@O +%# ]˽!aܽz{ͷvQh쩎]hGތ5ɇ*DzJDRNLi 4:{~2FmXY-zzĽ^f=]uū{/+&c:Ma{ĝDp2m܍kHș/(--m_vݮK(V{R}.k&yƴ7i^4@3f sK3^Ř˸B=]?gt5KbZB<e;kQLpxuWC}n 5ҴepB##~q= `x]KWF {GfŲ}?G.I9pjWkU]>={7q{kO/^I3==f1ɏ%nnʫ/Zu_yXN<57ۍ'vy/"8넭M2eԷ&Y,в33%IkjMr7xf nmQkX4踼>a-GcIeތw&U=-:qnW)z¥j :WqSZvԒ#j"KrIU)%qrmRoDGQ~SYRsu*V)  ,/x)MFD6O#]z 96[Ui(JRfw'y$GeUީkdMF-ݻ98F2d[o{Rn0n-xsV6Dh|Eb2E:KCOӪv4SJCr"J!!m,hRLD| ZYFm/X~ΧfrN&4Ƒ=Z9Mh.Mܵw/BdrܥniŪ8ɧ|y%œ[M=_tj?F!z5\evM:\ ~F-sg钬OWq“iiȍ<Gi%%n2rqͻllƑ)okw7}\Uk-:&fj솘XerV9yZuʼşdFC=rmo%~ZN78X(N)_7.Εn1MpJ}62jjJdI";R5&iLԸc:jmqiQj$ujp\{;v5B񥍪Xn Ą4qOERjzN(Ga٠䌡)p*v(J7#ZۻZ8O W uONb+^Qipv9GvֽƼϯrYƖKGJQDNPhRJjᡧC"21"9ѓS1;R_O7/WGz)8fE%F2ukmvSov/iZ&/]~KmI[:^~ͤ\kMi稜\ywJt3W7 8Ʒ~ݥeFgѼw"8VVSج\뻆}ݭ/J6Q)d|)zU3>k\L=;ow֯gN3pKѫ|wmkZ$z^2R:E)f>ς нd|#׆?\ǔpV{;\$ƵE%-ͪm0S6[n< kE[}mvE4DDZ^$OZ0*$~XUv҅B@^?]so#%ojw;Y#SxxueBگy v^i-)s)zV jC{7Gt.w3v,ygg8s]aE_,*E tY5k٨h=o"m泏:\6w噓aiL׎n^c\75AGkЯ0Lf46َ`egZ˓p/k;̛]kq!ݸzpԭG"}R9Ve>ˏHUjJ-&7nrnwG*Xv\˱/vN}O)ʼn&CV͍f̵]r\PMB-6Du-#RͰtRN^)mT _}nSȕC*_xBuTkJW[`ɩ`ejvsngP ڻ.-WUtܑqԹQj)t;vN&RNũT+8%IXӃ5fK՛-d9 ]CƑm|nZ-6=Hz,*aEm W3VzRšdY~Xf׀Xx"]s;)5u*ُHB BRGS6bݶؿ 9j[1*jױga7oX CUI%0v#~\-O-Ꙛuɷ쏪&5mY٦M`LJ2qK~HZbr =N'YobI. (^ ׾{_ ?OJ`S`3BN[}5w6:ǵ/iSlt=4F*d&T4y/#. ɵim5Uֲf 眕6Y7 fơ=3dϕq뚩$qTM-%r!$@A? ޾V0c~{[{;򥧅a~ڵ»&ڄv1ek=wb MLkNAԬw-x>~/r=e73VeVN)K%Sښe"+3uXuچrn ֺVzscJ峻m}vb㶓n\YbIUBT%*,0nov=;z꣓S/nSXSpl##k9mXGrZv^Gde!ŷRԠzQyjC]`gToPov{j~KRBMY}i[߶9KL2ԉO0K#m>wB[ٍ+n[[b٦DX ݲpo] [\m5qdT()mo4Oy9Ie b][wղmM~vmi۱~t \}$яimRk(L c Cvk7r9_r1 ;zv|F@KyZ[&jEji/"6$69ml#e]9s\{ScL}Ȣؿ0q/nZ*t,CLoD߉Njǚy=Pgmu6^]l-["çUʖMlʍp-"qmU>۷uFOJ%Ǔkx 'g=睋k[3u,{³WɘݪF]ՍeFX"Oy\,cچ=w/gn Ļ]#2? vqy-gXnR.^}ݺFs{ŝG]}e|#0mjx"ƬWكm?rgU^xVB":Dt>@LRbun~ݭ,w+v⪕;\U(RYa61>#Jm˞Μ9g9XKaG='u8gf}'qy#ɉw J]We.ʲ-<+&q%s?2dњztҼn`cΤmmqMdz O[-ߩӲ&;[tmܝVnr">{x<8U+p:Ig]zjGkt,uzf}dؠoJaکqEq -(:d<պ=eKy[˗^%ZXkX[C2߱\ITTLGzANM￵i]K>UsOGDDD.ZF6* ҃V Zhz{'xp^`wo8r0h ZmJ5"jb[l=yUu7-;7IT%:jFjߖm0tzU'K)څNۧYJ)4IQ}^KWm7kSP>q;ނ#)'n7&׊r?óM{IwR\j2Qn[v pe#/tAF\ϵ225q֒om6z})6҅*oqDsMf CNIN=T S2t,_ѧ}kveMF0J\Rnnݙܹy[rUc-j{yGtkQ%s]5qB.Nw.JN1LvR Ui5J ZESQԙr):MJ+g}χ!2;q([jAud][ljVK3$ײSJI=/|&tl'*n۽f.frܥ jQO8>&Z];.|7T/C}$ڋUmP2Reҭ8hFF\L 3~e v\۫]ݝNmrnB%*]Z«hKc=BTLG :V74$=Ǘy+EX'4tn(I:Ѝ;Df8c,k1%dJ6.j6ź{N~l6&*fœI7 WAlGOu-ҢH,,(ǔe뿋쩨kM܍ZſgRvQ' 9)?n|er˭|I|-fGK.rΛp8XV1%K6mvG+tc+qE&ǸC_Nm:l=_/m5^[dߌڇ.c<%:)tQ$Ow~-aY;UJ>=F)2[nk؆?훐M=l6[4(O.]2#-H^n#->&mp5~Fӛ+|| S,xag%qkEUzUgæBhߕP(7]kFnq?֖CpruZ6*rEڊtS|*tI*E}7R<,nUU֫^I7Q*mSly%rdȓd8hE<9oHhMfNSRj[i7D[Rj݊+kდq{"$$H?p\̅S?㭻;t~R߁)^/>Qj`yt[w ԛ;²~+ߔ_ YW~|o]?x^ᯛ `ʼn;g)T@vWn]>&4lp+$D̢1l|ȨF%-}.9[}w~ ԠLM9hСablfe&QoW!s?wjLK?s7yO>(=C~_nyǜu?v3vyo oI@qV-jeES^[9WoSܝh"l2C1a͔CiJ@3:Pճw=/7ovuk+\V;lDgն<[A+rX~d;m!_s8ݖ׷;;.0llUC+?i#_crʙ1~C.\–q ul8Hܶ2m`ܻM3Tov|Bs rɵ"oLS- DКw=Tv@f'6|YlD͓Y%׵-#Ѯo%:&!3o%\J<02;K87>^vgƓ# ;ݝmz^Y6=PS39U%~ &f# }o!muH;ʲŇ˷yvP+&.7e[3'vR4Yj̗IZ`e˽3o[WU{ m[sUbۋZǾۆl6~9'V*.\S2<Sd*zY[aŶ`]C$n.v^Ʌ dng>ەZ,Mmϑ :n6nϦezWqUJ4! ۇ4R! =>>Fn|Q[{pRO17ƕ~._I''00k=b՛o}Osðc2'o\3}ݭQ^2 . R1yKȣtAݿ-uܾw!`?1Whn|gzUo[ECWwjUIן)^h#1ɭ!/Z np;o;ΗŻkXs."6E`Z1 עӐ9Kl8qd q} 2Stt;#j>;խabONŗ=fwP1j)l6J̶|gV2`y/0E˛6+ԫ1? 6}KW c\KoKͨ2ۅFw–s*TԞLיuDx .kCzWXhy۶gLu|%TnupǺl-S* PRaLnT+c+*xl.v!.U=|; !_L̎뱚U=4hm:ٯ"y)$:>%(n}X'p[ȴ ^˒4kƓmzDx \ 'NqamP7nyN݅=j7%McSڵj%STy qXymvCg{w/w=wSW5r̹u erծˊsOm=DhEҚRb#n)QOxtվQwe]I}wCa'"[ۂ-z}2UuKP$㜉ԧ:mc<Ý>RoL?wu|%ҷ&K y_!y9 ??:tq3(UU-lkS'ɸ@jdzQˬR] EVPW1DJq2n:,c|ǻ̑;y{X,ۂ.u.b˕u.tKBjQ"[S園S`ٮdNبeJ&9Ơ ~0a(Vm٘L+Jr*vڑE( x0+tp˕ n';wm-ޜMOxX>{#2%jgb2M[`K*\5@8l'e=0u+w ֘鳾{y܀:R*Ya]"Ӧ%ktynlۣ65,3gU}{GYrb;ge'TKwǘ.,rpܚV]Tr,!dp /ԺU,xՉ>s׽~W5oTh yx?xrrx?)?ilbT׬,z$Ԏ.UH٠\U1pU:]JwSrGZq8àd驐,N67QYBӢD㏙W!Q25ϸo9ms-7-%3CihO.J鯽-;MZM8ku-7k9S$8]q2E(}bۏI[DKOK}3KUB^u %Y,u.-&f#]'܆o$x`Yu,dzwM;#oKxn;\[d7}Rb+*Y䛂ZuBӱl{j0O̓}LhK;[aֶaGL{Cb#S.T[>߃F]NK"u^LUʐ_ykW?!GRj29͖qa'0[npcDvV)qz9R)PۨM^aJx W] r>];eN3vxdmĘ(5W2K1䪖weF{mE/QP6\u54x5[hۮ-Nk”i[lUgL]J}5 S:EhiUrgHl!ŒJ$pe=q^b͵Q' ?6|R\,JA ڵ"TDꈭ:ymg`B5t%M] <N_zv2_Ortٵ/i/ReӮ*7[qүqEG* m"[I:6e^p"I$jԴęh!m)]GZkcjS!{e^z}+Cѥ9;R|/ֱeiUԏCNu2Zhcٗg$ݭwvr P8*7/Lk~I'Km1+MW%Bk|oOm>-#qj*|Dbѱkn|n{v#jĮqNpMIUm(7Liz;{ҜݞڝVƚVϬ+sO!OstGvxӉ']uӎ4g_ 1^-8ۦ k!)Ύ5O;YSB#2Zzχ;<.ֵOtge~.(RC#wFZeGZٸ6FFJ4e2ˇpJT$[wgV)q6muDGJ56q\I!̗ y/I~RtJ9kJ]Iy*'FN0s.[l!fw'y(7$œ WƫgyΙdMEU JQJv̋vmrۖ.jWR_M֨djYgSj0^\y'EoECjm$ IƩK>Z28J2TiJ2N#}.s cArl嫶nB.FIJ.)۔\ZiM>/hLĸ=C1s[?YMqp|94- 鮝𦔽/k^#NT(Y LS$6˩}{;5 )B۷W$qpN)qqoot}ZDVә;7TiK|6f3h$dԄ}fqݡ>Nb򗉉+ͶO]>ߡ_VtYf79ڰիF sq~prս|QM)g%l0ocJȨHz V;Bb/kLAcfPJ,ԭ{ƍgpjNR6VSI*$!yV足jᇑ.](EܣqM\qJ2eZT).<9UB/(B0j)mtKEj#׿fDI-=rZړj|'Nڤ]k*i$5qt"ݙPM6E4ke^Z8ۏhz$Q(R Ay2zfRñnpnkbkI:=j &ΝșW?׵d{+ύM'??XqeeĽ.[o=UxFS=ӷdZwenՄ]_X=ĭVa* pKs0ބۍfJ3 gz̚i|wnxtjc¼5${(1fXQ65ȼb̶Zkn>%FQMJXӡ{TZEVNᖣimT/37cNJUPnP҂ZOE~"-Rc4^b- FEͧtf5[)S!OZIښݲ͑;tvܡ+N)AR=hCNn;wL16-:特7M$=Tҕ-.R[HٷnXk sn[ҞD-0WS9p9:-Ϸ-jѬNu{ҹfv)[Ľvwfg(ٷfe+0mYj8Q1\ݧg]Eǎvڿc!4#j5̋C2"}BRriFp7=ô\TZ:\BLfj#I22װ<;صZl j 6:l"6]۸ K'6RTѯ^ئOԓV\?$x7s#r:Oh{ց=MmuHԷd{pN /܅:UE#Yy+(SgQ(Щ)RHzw>^Ѿݻ>mK&^ '$Jۻ&w%F|xfz%˳ L~3N?Cy9 v w/{ƿ kz3x> sXv}vP"@WyC z`'톽Dw%-tt yVY\wmuPYQA0iG-2JP,6/gˢ]u.-n!Zw.N7Q]Df}Q0({a\@=i_X7gFǘ8^⻲}G MZ1)WEfO12G+=-B@z\`||w6ċj߬m}UwRox֢I &c~XGP6Qndpvܻul'V7^FJt^{b^B(L~sѣ6@߿^xqU!ڙ5|Vpvef-uӥ^3  FSDɯKD%0r}FF穛r7 +o"V8tv̖NQU!5uFd"bCr^bJ=֤fM#ʳԷP0O-9xRBm\=`r-:;~3Tl(nXtXi%2Vٛ#vwqƴ`L@"H‹qW.j,JM5B[)WܺUeZFqc'V˷1W7V̾-MHФwn8N;HPSdݷC7&2j.W\τGŎ'Vb]c.x+Rx1%C2T{myg[qU|+m:M:շ8҉yWd)ՋWS%%:iqlʹmGwݹ WnNŤѩ5(9hTٵDdGUi-)vSs2 2{OnT$Xck n:¶(lASLeȔBjμPpTb2~N2~%^k[ܗ[Jzs0ӓHBKq[}JَA-$dFQgjxxFv4r/x*Rm% `4J(&iv7SkԲmSH1YWmx 8n.k']:Z˭_W >ڃXЩ. jTq%Aā[E}amc]D:rmHRiu:uӚӢ\p(5-q%e)(۬ҖȽIf<߽pr&ݫVfY91q2ĭEQgYbTGQ&,yL+N$[q*RVۉQ=FuTܻ>f>f㋳8N6$܌n)9&»iˤsX,݅܍ȩv+sRTpO}d?Wn/Inpȸ%O]StQO|v5\}7Zwb.AIVK^:wb{[uݯcytO߶S<{8KSRׁH̏N7ۚ[xkwYy_'ZӵF+>쌛ZUĦreE9F[24De{}@:ExWs-\ǻ7K-\JNvEk%:s˙#κ].oͳ;լ7wB6nwu:$L; DkI#Wz.:Xp(˅v$Sq,wn\qIN-e<5Oe+vuYTpcojUI_ާP8 O 7&VL8z$_B-H-[uh]T{|8=qVRN-:Ij:7PUtXϷmy鉿:RIM~33ӸS2#׳GdŲ5+/Bx{(WzȨ5Y㞎#|˖+ ط.|e<o/rߔX>7s}VE.OVti׽ .5nNJO"95{#q}Ay9do]R"M6z\tnNS-D!@3N_jicWsy*5uٮRcWv/.,j}=S)j5C^> Ie =gu9ӛqjtz]۪TMoߧI!Ǧ¶m:,"[L!{qAv-o 3{"KʼnrIkfٶj2ƙ؄S`7` k6jzޞ?e5G&6uʷ2%ԒRKE*G\Npom F/V |C0.q_eenƣ<5Oh'67ɪn[SĽ{ڔjǘzs;~׌(ۂ`ܢ1ƣ` _l9Va6%UQWh~P~\F^ZHR@:ۧCJ{ôGeBh;~ۧnU J\O+n2 RҠ)ng}Kh{5+S×ܛ.1ZjG)iRȤIN 4%{oΜ/eO[Nffd ĹK?nnԼMqX'܌nZvq<ķbFnͪaQ`5 s,M_լ?-@_{w{ӺձJ}GF[%v\5[ŒGkOw/ΜM9rjË%2+rd~+󲕛C9U۳r[aJǭm|˒LAʨSCq[XMۺoubfp:t+ΤĻo ][ zt-*67kvS7D·MMCQXm;)܎n_h%]4ܙnRk!]ڵsDUF"`R, &#R_*[z*ZqFXɻ]7|۵w+'pFDەs=r./ᐚm3Hשy yD"jHCr':sA65نѮ^o1V/ f;nFr3VM)e*- s D'H݅fӧ\*޷[k<7u<-]֍Q8R h|p=WlW3s%Q %3l}@U-K6f-NϿu|ڴmWN׮[׸F*mW\%r! C78:޳vBG7ŵ.JթԚ2x)ST!řn~9 W:Wpܢ件{xf8ٳwKE ҰWxVB\qBZ 2wMb[lGSnyԚ~z9ZmያvoN2Afnݽjf>)j3 !;gOYʹK" Wftڎ+׭b*2ϻK>ۢӱeyԪXISUm[z+ugX%0lϏnvg!;t{BqPj>PyvR7Cj]O%+ݲ :qiMj6W}3vC/R=4Som]ŗ=ю, TF6U_-\6MyskwMr&Q\wjKܩyMϣUj0*}RZܷSdY3>Zjqj6TgzpA/M`/Cmл,޻feE[/+uk^Vs1W$G(JsW2ٰu*߻q*Y޵.Wi:ur5T),=0uRmho.twܖiYwrWHntvEj8qhf`Ͻpf(R&>Ki%I7$QӖm-2 ~yߗQ-앑/ x[k8nw.c㩵k}]FkbJl:{.(˩n0Hqvαp7 귎.Gupx[N`Yq'+ruU7[ү+>!xrȫoSo]OC# d^Q]\>!ƛGw^Mx"-+%vdX-:M2UR%d>%l ioSu6lsj7D P>XxHz Ukà(n^Q V>5cVtWj SEiJdznyej[lE' 3kuٌNn4JW)gB {4 j6&]' m-(ZMEz8cz>WZ6#7+[,MR-Z!4ܓtCyE|umj1ƽvƷV\;%>Q :#Le(iVz5 4ũۤUWxX ^(ҔsլB2w-V ^R+; ˂M\z+Uwr+RWY⺧~ Q*JcYSNSλUd8in=v K낫k\IRרSUaCFmϿ5̗P|u ZTԕ}>oYѲ1sfP+sQkX8Gb~6r,s>^\,mGL+7[n-E\.Fqḕcl*Jmjb5 ,m]c}NXfeVlǸJ5eˡ$4%g~N p4Y*WwW٧<8v#;qԩTut,m"#Y D\5V`\\Lȋ];LȇiS6ϝZ l>LruR\v=ǘϔDg=ԈdFZ+M{=|,[;0>RiSi4,S5}yxw&(E7&fݙ4UՕ! ~'Id)]ǽu2K-fޭ \08Vڅ쓬=Vy^^ IhyKR-B#Ըr=]mܻӾ'*Umkoy rTqT_i,/8Q^<ݤ|4ԻO(܄"'5N~#m.(Ҿ2i6Uev&I*<}҄$eNtÛzyWJubW^iBW.܅Wڮg]irO6Ve90sgv.+sV޿aޔ[p?3q*FutUo*eL\KM'EG*ZcAFfG5J 5jj=MJ3OK:k˝'NMB7m3uFҕ\-Ywg%PRqMIyZGY9|μvn߻5cWݷa^+X֥vnݘ\v7m>Fgzv"-;Ew֝}1|RjN𿊀7g#֟*GQQ|#/bo]p$>_Un9гUbn9׃ErQBU-^vDmVh'<R[fdHT]*~}3j;nvjc7s-rӳ Y8[n[1pJx kX[Jk9Mn!_Nю6x:iZ˦U |߉^Ԛ݃hYxk &U^bwKk.[jE+P(˞=9j@snCv7%c_7=xǁ<l {t'酚+1F‹l׭:ݻILruǶkL-L(K0L1&>wXB(pm;1fpnlp֓%Skidkt(U +xulo'/ڕeN r=^pZZ:Pnj8Hf"48ijY[ N[yZٻ+=  ø:3 ?^ܷ^Sr#YK[UF?CuhC b]GM')mڏsNrܗI]ljq6VB. W,UK"YX5{c >Iqā> T:n!,5l2VzCl|+I[*SrjnS6٨y+x,@>П.g+!rn9>N|W>OZT_ut Y""v7|sfި;Pclm EùN,{'fNT%U&LfH8~1v>Il}统u6P˗c(WV~H^bMU.o*oOF0N:_:6Smr_.b+|ݶYY غF,mwjv>f*>QM뭱Sd:`N{l/⎱;n-z~"Gze퇎J5S KG9!Gn;N1 ݎ h6m|S?ɂ5'WOÞ 7|7^ao @mxGmi^jϽ>01Mf0լD3-2T. VXR"ɥV Kl J O7|u?bvа;6.eߓ|[1bmRr,eRz`z 6܎-ͨku͹Fː dPhYgZUj}nvX;z=gVեTv_J }\1n7w2J?ޘγc\E 1Aޑzq;\r]]\Y&[nsNei\uURje*Qk2CSl*xJz-xٶlm+|UjUؓ`Ladqiĩ!Gd\W~fz;Tn*PdRM&T4`չSWq5k훶(N"Ӎ% V]֦wb.nUO!u*J&Oӕ2e|Z=eV쫚΅g#+/RW:طnbi*Wyo)p{:ETKؚR(RY+r웓r(IF) VmȵNB:h Q1ғ|u8E]{,'$-TR[j49l*3"I鯴zhd>Q+\BkNF=.$ZR4Nwհ(IpNi.(Gi33#33e$FXK*NdWrud[r{xnk$v2ıh+J1TQ[#JQl[tRO]LHKٮ NӍnF񨔤֞Em'MILB"ԋ%dBŋ+p̿_17jzT~4pc Vo\ƹb9Rq-'1j;8ܗ)hE%DZKS<璸Bu*%*Yw5ڻ9ۣ^z4U; Ñk\U(o~G?VUĎ:?P?_F_Kߤ~ᓾI |pr.Ok\SklRhҪz{­P .}SktZ7UQ4ڌIM8̈eaӊJZ%FFZu,KZvln廐SNFIVtuNi?CM5]+Ph,{jN JSR$IS^tSUVrORYu.9WyP6 [Kiu m!X|]Y79ӄ)\ģ)pbڳr%*&ꑶ_-H*dzk)1 V3')UAϹٶWRxe'պn۫h7AR9 EAJeGLms!%D| A 5]/Q3eb̄vnVn%za\m kZnv([emqrIҕij|""><hjJשvvǕ|Pޟs}V~2&Z?+2N&Z4w@)4iSڪ_>/JN9Hiۏuf8'It[ȲR.hZ$ȋ_Y ~U<UUO*6b)Ovzڜj\R̋.$FsQuҊj^נ䈈y<zZIuP[}Qm=C?zN(Exqu/kn S-FzKZzOסӽjJ\)F3b!r5ٝ|;6 o=-3*λ]αb\abqRi-w޵⦪~b8Kpo)Z=>)ғ"5/GTZLE-輵f7ݘ۹~+&+w/7GFI:l33fg.N~۲\2|*cnermnnM+Fq"ѪIz%j =YW8@~gc/~?N'?)«qȸs➟n=k" X“m֮VreMh2[uݖ] *FܖN)MȐ`f0 g,C9̑o;ddudJ=In13:ݒvvdMUEJLp^,6t-@͐9'{7m{-3,>hnF;ѰM)->>+Ěz!R* :`e--m7nB\u{b U>[8֪]6^ߤLʦ\DFNo$$dͶlgno8OrsQ\l̯hRo8tuNo+ CTxu!2[>ctFpeޓƻֶR"3QrQuOѳgwQr;S~)6HhZw/GgVTmUf_yt7%$];zLWF̰xy2Ʉu!MCmš_0[W6jf#a-KLi+3Q7c^qg%s<1aYIQeZf+}>;S6L0]Yu_h9߻<ƅpmiM$AVvŚ,*#t2.8Y)-Zhshü97/#Oro"u^/uFgWɺ,p:6a,^x%$Yve^3PƗMnTP&yS}OJ '덫MH^:rXԴJۋ/rI;S*,+yz1hv)Qw^ڍJ2oL׊q(\fDj:^T%vOadɂnS}ZO)N*λdaȜkG_PIEO}нa(^iQX᯦-7^)%g'SJx(.S9zVɴZ{E ))ۅi/s7 VIV-|sj0*UBTHIqRf>FP$KqN0 R̻8j\GcC}IUz\i 6F)Q{Gҧ3qSzKj-Az VЛS-zy:8*mNk|D鿓ND2u+0Yŝ7kqm·?8Ib]u>˗^_>(]vӋzv+ݩ){vZrJ2RQ몋C$z [,pp,8mڊbR]Il .f~d/ݓs㓓mͶ{mgjQwn=Oic9ܚm4Q/6ݨ[TƧ?nԶoytf{@AzT{e{[O'ZRZt~AGD?s3􌿂ՉIw'|~U\ w~di:Kޱ)U/sU%njѩ&GSP^ǝd)..!^U` 1wX[aԇSxoFV6_扐)T 2Mfd=ۖͭiZ7KK Bi9%7@<3<ճԻU,},a}FRqɛr i@ONJvK KLN M, ʖv0n-]DwlI-X6ܶ$Jʴh5O+mOI+Ra瞠\ MG7BفjYo1#͖0V`Ѱ2M?c8>-Crt*JkIGS:e#hPKx[鱼>{5m;wcն&>j-M֥^َ) 6yȜl_w{-ō̱r> U=]iw3)r*]:K]6BdCTZ|>gf}LW}[$'Y5 &c -j.z6R 67MԷFMnÌwI7w5E}o޽+K ֵy4܌ȥW"COyR[q5Ӱ͙f[v"_#q{MV6܍3"u9BK(41ӯqˇc${ߝCi6I(OmθzҜ5k^:>Jzw.>qV8{vU[ڶEm|DžBz].KHjI]x;Mɗ{m,qZXr忇2u^RO2Z}ZێS[2Jen!*NDcrBUً4<ǼMҲs1Zw57c3&ĖڻzmP*FuJG1-dN:|OU}ҵgi2t~F^^Z.VxjvŧnNNh<:]^~NN+ge^g.SԔGFe߯'[vn'(ScJ]kܗ7eJOlRrfziݮq̋S"\*U<*W]k$FջV}? 7g#֟*GQQ|#/bo]p$>_Un9;l S VvQU%OLU{οmU6bZ1MTx%!֙Q7, J=!3 ;Q,ڌ;6ͱ݅q^&ߔ·n #WbwӖX.HtG)N&d̵zpI,n cu ޖUj+VXUp[w]N o.J6Z8Ts&utxln;~HPHS/xw`G\ʡ¿rj Z^vt"[L:SD\h0sUwR,}[x^X,R2Vn< ]2YDr[SRKs8tXb̷G?Ps Tv 3be,zVz D[/I.KOEQrm'$7|[J>r S`5յwT#\w1FTz\Ԛ &"ׅhSHrD\'r]~/>p;:Piuu:"9ő=tTaS7V2rӷk7mb[^WmPp*[y.Þ6f]cizJCgRR@UVl큝.WJP1N{/\whZ ػϧӱE7|E֫Sί.x-Y&pi%v''-x6r'Ws*6=DwwUu]=C?MK [yrtܒG$!WGqJ*%SAz ED[^)/tė/g=#Omd.|^n/sl׉g DZqemqowݮRzUܜ=ڽ-o/Iۖ;qVʘgPp|mm;6zGl9.8pwWgsJ2qPbe}}UpNjٯ}7TMQKrؽtEx%v w߾8%|j;~|}pK]ơ/ w߾8%|j;~|}pK]ơ/ w&~e_H 8PL7:%ʭ5Kw&U2vwR_+rm'}C7#rWoO&HoG?M$UR7{FU]u ;# !Wk`|W>׹潇9Vn)6)*ҹ{%qV4q>W1vi#T"Qk&GwxcJBJ- Ϸ^ˁxkU}ԣ/3.;]J=<*)cS)ROK9H=,r zX @)cS)Da^ԽQ gxJI=w֣gf*TRj

National Institute of Crop Science, Rural Development Administration (RDA), Suwon 16429, Republic of KoreaJFIFddDuckydqhttp://ns.adobe.com/xap/1.0/ Adobed     ! 1AQa"q 2#w8B36v7XRr$9bCt%u&Ws'(xy4T5fH  !1AQaq"2B Rbr#u67Ѳ3sTt5v8Sc$4ĂCÔ%UӅFV ?_Aנj- H>>,m*>fzp"TrKkr^r.|_&]|*vPuܶvoQ1mwVJUhu-I"=LniAƕ8"۲ k*ҿ[yu:.vUQ+)%F DHyVBk>Hy8jݹ q~9D4KRmzQ)^ʔ.J%k_tVi5NTjg!'ky|5asOȻ)R۸ߩFMԿ3L4j6dڜ#NIwUF]JqB/(FafJRzq3\G՛ ?~\ 6)6W4m[O^L0E&rRMض*C .]Unl-1 1r#Rj/&QɈ׉˩s6Rj=5Tg.y.·Pӡ:JJS:C8-2u]d&vUz;7p9 5VnL֢"y)">iי(IDDd| Yj0; LRfS:ktYK%*N2^m|&dğth":ey)uPQZW)gcC3Pv&MMWd&Ŵ۲mvTRoժM03*F3Yd6\8,\hݻ kߔi<k NTwSԪmljj[>->ptU%'LR>&EBH$MQAUx[$Z6vi&_a.KIQ{hyƒ j"JOC9eFҝfj;˚Ω<[3_m% lQ@4g=5$(J]Yc-OMq<Ǎ wSzڗ)k$7VIP붾ͯnV+卵*t]iЎD31~SA1éC2u)ʼnQn-Uoi3:grI8ؓWm*G zܕ)ZקJ}Y YlGeJ6cB2I NS3Q>k=KTBT]W6+SOXQgGR? telˊ%-Re\hѯ2TF"C/OJΩ6r[N.0{SpljjX1“jOsӥ;ҭhe}xu`Ք&.)yO̒ Fߑ.$Qw;9Iw2o+RVJMSOj[SoҌZ%;`d$blQ{Ro{Imڌ>3egf\O֝Uzx"䢸g+mv%Gʆ:|V[N'&ס-ޝ'kfE|K,G&˳98Juin/\\Qݿ̋v~Ǩ!rtWU d|E߫R4d}.qPw*Ӭv5YEcn~f5c%MTMkb-F>5JT,})QHg%{("ӔȸWMsYyWNRrkkJr0XドnͫT}r-jj,Ŕʍ\Q2Ri>v$5!]"JB2WɅ)]VԜUc8i|.jeRO6^V.¸ Q&#|ܶ-*uOG%JAtRZRr]FFG\۩w+?'zչSѧt jz>KW&ot{7P&2D;&\\>Q2JzܗAKSfeNn[jRrԕf6,q,F1tRfԗ>vֶևj-&R'Zi2=xv~Elbsvm8=ӛ"ū񕜈BȩlWau[]ٷBߨF~J!|Ipr3R̴#Yp)={7:G{+:\W}n|Q#%)7^-h"Ƒq:M*%J&$T軨I333׎g_- ucBwwjp[6i25$̏bU’ٱRv?G\~#Iͪb7<<}Ezt" q_Inw,7-d,G÷%T* Wg1"䥱kq/A.,_KhqŒxwvo u2ۥۧ.bQ}XκA$֣ +K״ZUNmڸII{.v{5z5ѮRme[moyƾd~cRݾK'j.\i&/S6f|b=5: p!6i_ 4j6=.si˧eƾtS^c.Y^RJVS-Vi3,esi08?H$GvZgg?gi䤟2adw릿:"۪lkSN>q-4kI܋ێe̊qۅgDoѨ9; #T.Q;7#~_Ufstb_'w~Xw1Xk,vcOt._}v}8"(4Z\ۘgk?J?bm_c!g{HZV]Fkk%~gEt)b秴vΰB|꽸}mp~E6ݹv;7P٤v+ri*3Ԣ|'O14_~7nP{7ZU\Vű[ +7󖱅o#:ǥŬ\|3r%TJX]V7ez¨Y]lc|O3V! R zbJ'PnGqVJ"19WVeOF埜EaEJωqCN5Z g-9[S<$sUK5b|7sn\7x qmv##FF\ w[=-43$^ooVSiXօv7iB۴yg>]Vf"r$J3""32!Zh[K%7GvNLs+4nB/B{vlsobJaҺJR:0g%&zR\ S3T[&ִor*ⷳc3ʊO[iozW٨%$gn:ܶWwFBԹjHP&z u&F2\f;ipW73 [; '_̽b;vib!oec dC-tS__$Xs]l9&z$2/N>%'[}b{h/{`{Ji׉׏ YJB/X%}.|+{(S:qz]4_Kѵo`^tY_4S#* ^zvݾMr+TrkQ g.8Ͽ^i>ӈǙvix>$o( ^qt*&t1oJVu-ql5U6jCЉmĻ*"?JT=K'O/|=Vo}l0b}}f?X[?/\JSBe,kP8ETJ==?.p5ފgbU9}ǶdNKk—_$8̸͓ۍ8Di\BԿ-1v{FF]|.^ۅ{vl12׏z7-R7wE?\nh\jN/Kձr_oBw"N QMBZqe-m:ӨSn6j4%!hQ;sv'm4kcM=!8\m[M4{SMliۇ%eֽR&N:{2A8)THLK3Zj[jPBx#BگMf:G1\`edcʮ?|w(-̮vXt,bW2;.ιNHRR#YwTM"<;mk\.foIDjmlJ;vxy7o7i\,KQŊ9d^Mmgc L*.T6tLeIuOH3SJQ3=F/ʿ<9\JM6mN6=<{xkP!F1QR[I$6ُimXu2An2yԒMU q f[IB-'䤯jYm52&JG\zд\~vdg QtHGXw&1Lw+nDEdC1w|YJmvP)HZ>i0BPβә?R:QO["]I_Jʏۍ>QKyu^bycBq4lXF~l [\*N>-J6,Gq(Zr5h]CwYӤU~ʶߑ u*SIv%ZfJ7)! FS*s_\|IŸZ)J ]ܜi4"z[+Z,MOZ))}|Ʀ(RUNIII.S'ˍO~˨rn}M)xxӕ0 eyҵ7YMAB]ӣU:/ѭ*6bcwP͵ "+qēVjŹO|GtY4V j[mLV M -m>",B$ GD1~j6O4|LxnNmqATNR3ε|DŽa[fmn-ڭ+FiK7Pcm;r5 l8r{#-]'nrFh2ruycb;pW=njRqRJ(d mnpckNnʹ+6]tz~E=ʕ l ZZ5jSi3#47.Lcfe`9؏v囜.F\-UZ:*0_<Νu9Lӵm&)_3\^ҹ3"1n1v_|uRʞͫr'iȧN_kH׺8xXrj=\МH)V\ˬ.Xʸ oVRC}ySU9/OBY먌5 ٿwޞ)rw8Ӫi5*5ZΗcGƱ !ZۄlmpjJ -l <R̵/JAպZuq\IdUS 48wXJJtcg4cI~aqߓwŷrm-v)G7yS^7H^-\mŌAq|"m9IBnF㏉9[N+mmy/!KKۉ%n +BdddfFF6FQRN-U5;Sv'm4kcM=Mn)\qιqUd9F%",6MGdT%-+~ f%+y֛^3SrF>6lc(֪vۊN;g._0Sѧ]ETWرkQKzGe9ʨsKA"yC y2\[5 rԭ7Gk5Mzw_4sM3hxЊ'oÍ5jsub )ͪ~tR2H]R͍>̋m6=%(˿(Wrr-܅y5(ܔJ޺YunW̹븹NsqK ]/QR#"ZMDfD|43Qw|._ԡSqTZBg??O Ϥ)/E_U|i}2 9Z?¹0:x'3,whǣ?C y-A~=daJј&M?D1_PS+Oi&;a @;Dž7[ zZC"bv:jjMQk$M RԸ3uA\=wI.AwC"^.{?-\NSiˏ"b}T/}q/ o.1M}R%:-ZniʒL$SgrBW*,Mw'N\ɇ{s\j]VryG'8f`}'N<*/`U숻z CwHq18J+vԕKss4R53/&XTt1bZƟo\=%nO)h$rBi-nKĪ^ ջڜlwkYm[̑+/QrZo%TQ;TLs($2C:s.%+eoNttq۰kK7O0m_t_pZ1SsSM7"mevFZ[w -FJ*T*jФQRg BSu|]g:ɵzjqwmltL.e3sRMچkSmjkmWœިm++¦'tILk*բQ D,PB\lI[9{%Gb R6öۍmX-MaʉA931cs..G4CujQտ[9 }G-xwl)IQz j Ó"rqe&=]꾧֎c)<kӳ+0JrRR3'TnXi^xMF Bު*tIL.[h"2"nKzZe'ZV/RrNYz]8죝n]Ķܩ>^Ժ]u-7^\mZjܣ9+Rmn ߑv?oꋘ?&ƪy^N4o=3-ؔ̿*`}V݁ ƒPu8%$ ݗ]wt;\y\>='OjPIp/nJU8{϶FNMsf"ίNqƹ(+ ݮF2Km |jܴZs%zf*eȫ?]4)I۵nR&FX + [jDh(#哑9q9Eծj8noǕZf\J-l&Z˫}`ӎhyrΉn\űn]9pʌӣ"׮Wt?N4_I_~54#/my1Xr*척aS#DT >q ssΛW;3oUaJSRMDgQnt:Ql,/ ܷfRqiM Ȼ>Cob;A>ڦWقM9X~/!'MW.}Vrߔꔵ!5|iB(0-zF=}okڢE$^wW~nokY߮\6՜̌{i-AF*9)\t9IV6۸5ZUF6R$ŨQIq砳YUZ]eyv >hI櫥N )&l JulwE1GDOuFN2| }馥uC1rޫV+^gdb&W[4<^e4YW,d|htͮsUM)۸8:{3d{AѢ)~ \#J=NdƮꮓ90 |1K$v*?мS ]i$J,C,SG?/_՜pMSƯM|mG1V1$~K>CSvkuj=&) -,yLjuFHK{c駗.SOua;BrSqj-ۍZ#'Jys7[g2z/.u4+XV2VQ.ޕ)$"(%)#Z7suZ%j }BǬݕe)Jvz8zJf:hIN|svO1O#IEcۍjݽ:SdὮvu^@:o^5cs>i/VqmVm]ؔܢn6'vޑ̗J4Wn@OlKbX ;n:hgJ9ŻyǑz8f܌q&Y fN0N;[69 rbׅC2/#kE l&2~èMR.*%g=Ft.%؝e8<.e=Uv{~㻏"EˑnvDѭ͜Lu3u0:U֝$[M5<:oi+V4V9 6nXvx&_ q Qqw3W:uϔ2yb/(ɳ|5zQiJ#r|Hw#.W?4aDŲ\ugWG;Cw鐢K|xg)##=O.dF˟jMUvWĻsr.z]kPc9"]R)mkfOd*uYf١RsB Aîh=k]ʳUrrZsq`d#r$/Ը3o^&lRWȍyuW̦Y4QDUMJ65ƒ[+ygk XK_±k#y:8(TJOSQhJt2.DR}"5[) r)6V6u5k:eXZmv𭤔!푊Q[qQ}ҹLE- 8qIZG|UM4j}Mܕ[Vwm{} Naqµ"ԈM zOpKѰ?IAD3Ir0'/q1itoB5{%wkOBn-ۜduqIzYK60{+DʕܞqIt";r1mG/\/ym[6JƫR \L=S=OT@Ix[TMm{>ݾտ֒ݸӉLYIx>+"JVNzx||5rI?C{oz8۹e\R-^\A2F R+N9 vlT]"ۭ d)t֞i #E2jB@׵=#/N+!ĕhx}I!cM`ąZ*ŻɄҒ߮Y.Z}='/oۙ3IpW̮hT7cTSuz9>B}΄&h!>lӵn~j˅IvU.'v'CSZw8QK3G> ,J59ٷ+HSg䧎hJdzvwv-cvxS5[̊n~ؿ%ַX?O0\6ne 6kn9.ϯ} *h 8_QhLݣ7q +=XBҲ5?[[)+F`=4 }B,sNg==u*Nj9k_GJ)+R~GSPBȒZ:(K]heL=vKPӢwq(NrG^ثϣ?#tC?.ͼ[ۅo؞y#%ǛjVyLSw%T*s92JTM%"YkQО.q)gCͲn8cgi6j1MѾ[{9h^vƘǚםidfi.^RHmg&rׇz:}݃}xT$ضk'5s-狶,\vpbPD،=Okf.c#cdz2FK5T!&)|ntD<+OŹU i-G[EE*FDfeaf2QƤM\UG_{ǹm%\yrGy:.\4wjPGUJޕUV7Do\7Vy_13w;[?c]H\$IJ,*L]3b%L{y.JRKG2sq,B6T}(#nW|km+q5] r㪍bJ@y{byz,b踊3ϻJ,'^xd،)JVw#.Vټc''ÝպWtbRؒJz۠8!o9IۄS95E9ؔ-e9JR{dmnｯ<[~n${~Њ$W?&ՐY_? #a.ߑv?oꋘ?&ơ|y^N4o=3t=~7!/M3>n8W홎2M`Qx+ z qy8%]7_~540ۦ彷]Wq CѡwkďyF5Dum_}~P(5.(X,K9vᯐ?leB9;Jhm#3{CxGE-S{;@Fz˙]=O'!ɿ]' r`:7'2bЖ>Iy,/eTy/V<.H?UYY{\^#ѣr9^7?xoRȆ7EoS_&??zϾM?(~Q-K&>"~aߨ t7Emsϛ+?;fCr)fY+>z$tIkjn_>vnrֳki-˹l= t;'EyC¥|/BLwBJdgjۛ$s S1|ɍV%JI6KvəhzIlBYɒ|0"Sy0F>eo5W)O+X˻u';v)2vVq۳kۮws?UʑBǴYO漪e2MIjPAک\b1)DDؚKm6ZWΨgȕ۶yjڳ 2ضN[C[|r@9Jfo<_eI7q.|cÊV߷:i.:$ȋ)1%%)ADZCEBxJ0MJۥy(bNsKM9k43IwNt.\%N簤I'.j|ƃ2$grBEٌ\}9:v*!n7M(ɽ]7c@XxƱԨ37īf62cTTfFK]9wntQHͮvٱI/f|j=7}\_V5U^+:uljSȃY(XI.ȱmo1甅jڎIZ2>#\*:gY|4k\8ZwSqtyA!+];бޞKծË¥e)#5ap.QK^8VdU{*ѽL\=qmjnB5>{ Ӟ`v±5 ^k&O~Oshɷ,;6nOW>u6{RqS`)S%jp\ipdEBLfTWy$GIYw~䲭J.1vSY5z.V>^+Ǎvc.I[R{QsNR3ӎfhd>y?UJ*}~[e\i5U^͛E]G_FS(Iɿ]i8:4zj~շsW,ˆsy:%O}iur]iF5~3M:Ӟ#N06)4ߧgdawIotiz:1r5YDZLHBSi;NQc44la=Y kQIT*ըl:tq2(է9VO4뒳܂~2rq'nrVZŦ[t7\oլfb/mlpc.I8콚q^1iE~䰳mi[dۧw֤ICfdFeCsg:i| 6擣׋* 96lust^{%99UNRvaMܽo ammi$em4D6DD\nA%$$#}۷/ݕr99JMն[oT޲E"KTaP+HGkŴj5TM5xƱOS-k`ۛkٝWz;{kS}F;~q|~^_|euwnE'pSupUP)V]vE+t =ZRaVdG6= *.ϼnj9:UɷbېmF_tޫgHjVS'śǕًdkkѻ_]Kv?nT>)^e=Ar1'3ԔILyD?:-^in):{7.؂\.:V }#뺾.3r̸*xbFM aȵz 6SQ:ײj[ 8nn iFMw rR"5M5I旘35f^j='j:nNW.ʭocZvZKV^ɚJ.cM1ZI7E'6rg탸5oZ=[m Z`\hbMUR١Ȗĉ):Jin!_7Dй+f̷eKҷvͨBPR(V`y6tw*MRΝcB.ڭTnc;P$8nFvm4(D(R#R-L -2:FP lxZKQc6I("Km%$E, 78uXIFA$RQI$JbInG]c[ֹ:ZM+n^')JmJMJRu{e)7jQDw~%yQl}BZujSSf۩QZ+Dzhd5o%BIc'GZ?}΍:>Ɵivז-%݌J5MqGWTVʦh݇ܟ~Օ_6 n'{3~mϬj'J11OȻn߃r Qr\3y٘+WӍ'WxEs^O3 o~[|7>]]H9݇ZomT@]?5B:Z߂'`V_+/MSKX߆ޠk3?o7y:4R/7þ] iG߬aBRU&?r&/} cQߥGj2?C5Yśe7hU=?+ x龳f-܈czW^7p%-(\D4h{UK&ӡn^m]Fݢ:`δvj俜F+) y[{{ 7 tu>gvrěOj'5 iRg[ͶFjGe n~qT$ci ۚ0oԹc*jL[sVWqj\ݻ&6"WoK:cnWmrv)o>66(F>=W^bf#c zzʞtپy%mՉPël e}J.\Zk4ttt>oEM=q)hJjI=ͥ(%]脼_88ф;͛gWG;Cw~˘$4=uWdĜTثNDkiQL9U*O"4XP`02,Ge-k5$h>ܼ]3vr6!9RQPIVSnM(ۓ{>;/Qͱv{3&-[rc)ܚI$n{Sv3[j00)-D3z}MRzVQпj,T[uVs0\}Sid;r(ݝJ>æʺL&c[jPK0~d(FKÝW\m]GTcF|Iׁ)I3~#oX%vҦEݑؼ5Żv2qAZTE^..M{ʐfȏ2##.R}*KʛZz^ӞN*lPťLf\G6[WVQquV]XAi)5J!,$iJ6o$tPZc;Kjx_n3`qIelV~vLy{fn匋Ѿn%;zV.n'-ұdd2߽1bZksPe3TI9)$ԩIN9Vơ\=2885N\ p)/a柛w9g_lױo8ݷ iixJV& ғRi{N^_oAŮE6Y7I$Nk$|Q)-*4Z)^¸%4Qm [I%.c-OV+C֧R#%ѨCe3i;w$G+_dy| Fzj$DI(=OA gj%v/]8qԯNIS*֩',Q%\44ZZ%D|Ǧʴ6&vֵI$%8(ԬƾS&#Z. }6z?b/|Jl{ץv&mpx4Z$”ڝ4-H%dGKfM:sKSRWeJAn]>s6应-W9'H]'uȫYvgK^\czp|My\鏩w/ËQ.)]\QiS`8uL뚛̸=J"ܻi\å'-)54Ue]:K\퓡vK xwBqrH\*֕TnzC.mT=t-H]SČ~Nu╏NÅ3f|͡G~B+Xm[Q7U{9"~jgK Zoʰ7"qJ,ekSeNGgϳ] ^.6:s}_,%eRg<5⿨z{ZPun#jRІ.6g T.!]xa c#jN$Zpl̋H WZu8WmMRýsĮ?Mco~sx TU҆Q :KDG4n42.<3/'^?6/ܠڒ^yrrÿr2\D}}B]^E~^T cɛ7϶Y[<֞[7d}2%QPqOLEQR\CIsj1?\}%tJ0e~ *sk"*)&ۓEi#{1J8Hrt|'ܝRr8)=ƔN'RVz:cf]F7bZyZUȘ4x8,#JG̒?.W9XnO]KO]%]ƻ O5Γ/3qÓj؍/r̺rƵ 5\&m6h.xoeX[=<3%< lZ"2h\Z[&jW3ejm?k&[]ųj+{N{66leu_+lj]q* 7g*knأYv= q ەdxЬZ|%GUrQ3jLŒqET]1% qkXYūYc[7Ś]QY\jko\</Lc7+'hMSUc6qXyؙ~6#ѯv.0$BQi5YyIhɍiy=KD!n3Vm[V%W-B%swa97ajۗ m+9~]fKq|Ddaˑ0A]_v޺mM5* F-BYHJ5}q>ʉ.6hyDmpD׬'-_v5;5[8K[viJ.3dR:oYHHh9I7:۽fi+wm^ [)odPѱ52CZUJicSw\&_s0uBȍh32džzQflcd^m|7GѹE!fO5]]H9݇ZomT@]?5B:Z߂'`V_+/MSKX߆ޠk3?o7y:4R/7þ] iG߬aBRU&?r&/} cQߥGj2?C5Yśe7hU=?+ x龳f-܈czW^7p%5|Y:SJE\U-(a_cƣUǽXXKiȞNlmۊڭڄR!**ܤMeȽ$|X5(Ź\rJ~ܮ]>'HB0cp XFr_c?f?7<ukSgov¥iG>>䙗i.+t+bOjIܶ . i^:nm}s}(3>NZ$2Qg([".>i.ƾ)B̋M8+"- >eE6DݥJnJˣt׻ 5.˅nJGwZD~!i۶a,Db3ZQ3O#KO5/֍ozuK'GbRi᝘NV_ҝcvם ZoX}F6z 7e5_e:ۓj=AB+iܔERadMBq*ԯ DwI/Gy*mĥiRKg6skY/#SN4e$-yXM YL?^ĸNNӪ{$r1JJRSLO]Aqm>V/s[~i/j+m>z}eI"Qvp]{ZԼ:{vPAG2=T͡@ڐ#u"E*>C;o$~C#_d/HBq^YRٽzIKbOm\~żjFFGdiQ(*/i*#.FF]©m=BmpQQQSP&Ҫ!T&^>:y)$ˑÐFčI Bӡ-t!bM WҦŶ'UZ=}zvn~oT/\ǒ'nr8 AJIӆz<^uߖ4eFC1i+v!3qNyߕni?4JZlmYFXFۼO0B\m[ tʄU3s"Sr(NJ;SKW72L4̏BVdf^Ҹj\]ȱ۪(ӷm?J-KEmWڽ^4<8qu%9pŹW~877ܾeVгS(յe^C]yX͹! םm4FGȋ\y'Z FX7e)|Gjt߹#gb\ŧq_([R8[qU$Z (ʻezV2V!iQ,i$JE˂٩ a(GK'O{vnBvryRd-RK4=qxZJMl_CuuIz @Rt㮽޳!|68\-l[џ84-2Pu" RJ_^OL>G1~XnBŬw6J0*Uvlږ1N G1q9IUm*'oWu][&UyYZbBZRZNfEJf"+2nF~Eû7n1xv.RUM$6 lAxSQJ&n5ܞwlEói"#>4׿Q.nEq7Oko[1wg8ZQwZYiqtm&~">Bo?w͡ni2峋NCEy Ҕ+%ZJ ʩq*fpˤl,~^Mχk1+:ݕ z&Y`KLӪУDr3[*Z :(SL&ݻ۬Vqsyԭs x|iI߽zZrg.:mp%6ԜvgmpIUt;QbS.Է) ǨKSV,*lڌ|5Jt3#NP.=+OZ~/G سIgbꥹJnl_DUM\iM!֔wVZuԺ,yV.Q>f v:݇WiaŸN5Ҕ[M7SsrvǣrMW= \8ZW-jsnڕ.ZnF2qt ً[ٻޘY۷Zm"Jxr&NAfA-݌to9s359݆mZ+N1-qS$D=17 x׵+%_ ve4ir6Z$FDڗnFtOr'7'{9C˨ꤡaYoace{Refnft RR"4%ʌm:Sj3)OdInTO>X'vxV#jܮw9Fog;5.~Y5\~18YQܹvj4+~t7S ﬕs %^۵ڴDZV69R^Y+rj$ԇoJKR5wB9C>Y:l+EǎS{ʲ{T6Wi* ^^9k/y/Cs\g*qڵgn4T8mERr|Ti+iPe;;.i\EBEJ 丬i9ɧM-ԼsGDrZ>r#R>~X9y4b棇9JwV۔%m(b[Tjvl}۩~nDԺ{Zo-YuK1vx.nWuO+jN [ٮ0%"΢CdTJK-RަH"$I(*ve &҉FzB,_Vpqp9m8werv')E;o&QE׵^d9˦j\_,ڵugZȻ̧8k+jK{wmr@3ӭ2 wFkzFVqs1؛.v'I%$[iT]D5Dl2 nk7qUxԫLS+sا3/ΖeZYK<["%-g/kRs:f3;*E ت wJ%)5&+&rw*霣i|sMҴ|;R+fm䡩.!**dӶ-6s6,]zAXMWjmnz%SJߴm2UXw7MQ%<!tKys#P,W>s;3IYwx<+i_\\\U6 u7P|xbn_k&ӓVOe䦒 VUr,-㘘"-LZeOSҠթrEvq8Kf%5%&K"#%vD/.ZYYŏ+p$nZkvއuW9㓱Z G wYIFyf)?ƎUm5ԉ/'k84{KO:rQI}XRuԪ|*lu)3qZ[mSm5R3".Xcَ5c®ࢫI*۳~wRϿQWޝ(EJrri&ۥ^ʶ齲Im|[yb;mnm֩uiܘq>E+Ikx߄3r33-5𹻖09ϖ9[Tz~mr5NsWl$oPusޛ^{Z;);sڹf\3oٹZmԉ/'k84{NO:rQIBø8Bݱ3n֤DiK4u& ofSȒܩx<˘|N0Fչ]qsp"}! QWw@t4ӭ+cO5%]'*{eM߲DRO1y*q8w++e!c߶ܪlZWّM欼 CQ̼빶lX{vib/V/ ai;x6~]+z]MWB>re-:lgk}պ!#9?%܋V-c[z!W?c7YNm/jRr[HOzԻefճ0q15Zp#rkQQ0tU-AmڵP/cȕ?0cZYj;:0ZM=D6g ?'UN+ձ[K ܖB2'xq9{|۫N0ku 7xaj;n\ 2[VznMlWiKbSk))f..)Km)&bGZ=>OR܍W:j'rM'wYz&/鶧{Sʵb"vջq[I-ՌZH._x*BagC'T(Q:$ͳQcMCKy?3g'ߝqnT);qs #ؤZ}OOI:cfnc8W~qy.;^pVl]Hԓ>^H^@7-AA܃nmL(uWܻS߿ Td95Bdh4t6*dDh!EhI[iŨ\L.&Nc ܮf^;$R)\rip9I|ٺ?#R.ZDZ;/]nݻqs\QE9M&Bd ]N mN*D>tgbK>+ˏ.!23]BȔR1ɝ^j'k2ƮqBQq[$di]icV/e`޵B.FIIJqbi>Ӥ|p; 6${)RU>_e}^dzdfzi %ekRVUS?6'hׂ)5.\+qUgzE2C˷ecŏ^֔ibk shesFWJ#~> Wk~ݨ}ڶ>ơǚ)׽ZƉo~B-ڼrvoE:Ʃ3ۣK7+Y`WirS):{>ڛ}:wԨ(J_";6R%[u&ƫdZ_\'np| RJwNeTW,=rrbnkڄ[M3ܴz)3- R.?:okۼ0TU'w{6&w7j1z3ON'fGoO?)S_bQ_¿R(^ԴԴG.EtMڇ&RUiW uQjU> Kiu1d<ѥIQ'RQ1:O/lŗᏩiʂv&Jc{D5 Tt)1.n[n۶X}RjqnOʽ(~[Ns{ސ⛌uO,kgo֢dRNQȄ .'6W!׌P朼tdZjFGE"]K@'i۪N;sI[{SOzk>`rRR+!σj8&TjlvA̷Q?HyjyLHNտJMjܶT۽lG?SnKN%<‘ nq[N0Sq[Ta(&t(|HGO~gvkݻTR4&Z$#ViOY1r$6YF?e4U/Mvxų:zbU^gQQ+NW_'4jfz^c'#`rvrڡ(IJ/J ݦ6 ]-CW |_{v*_q3^DZ}Ic6Uڌ8p7{crZq5ki`)mU6|-Z5^iEz3P=:Cu7DF'k%}<C-޹ֲ̱#\,(f88%X-N(ck0VLR~} G"-8ӏ/ϰKq?(#nrVTmZ;zióM4 m |UT'C^_1X.gXM{%ʤd 4\ovN":"y-,T)fLQgۢr=/CƹǨJVr[a+!rT|%Y\ٱzsS>jͱ.oOc6f$q% ǒGo;n[];ߎjrk{~\VۓNIGn:iqxo |~t5)Rxעri{Vi&NUOl_ѮMfsޕkЄay.0P{7N((BaIP$ K"U6Gl ݙqJRu+qN$ m#*p<|{:>-Ev=86N*MM긭U*uѾ?/^o7;'u,h4݌xښRM:5.(/ \իU.{F^rmF-Jɷ.>Q"[4xT^OZ~mK}T0ݛ^SAo9u?lX(' qj%=X}"^e4wˠ|rܫ 6I\Ķ;Ӻw!'ڍWg{ i U_9Avhۣƾ+:vs/MK[ɭīe{`Zgb}r[i'GE2J7Nez579wRq+Un ]J.cJ4M:h箽Wxxm^ pc\wcN%'My $$| :$Fqɏ¾^қP9J6Wxvu}ݵP>Z'FFdg"-; [¢cmWkÎT8nG%ݣ7*\խCLRYZͤiD&J#'ehbSyXK|y*ӞpS̍R`[pTr/Eg)K+92{_ n3zwz'oŸۤ+sOj J:`T>Cf*lwd\fYOP"R E֢̔L4ɥ :;.b(B02rJ蠟9>V'9M%)IqnhP<%,r'P/vNSwr#w"ݨaqc(|{kd=^0jTMR2ULNz|.<|^PfY22##!,K~E BEJۜ&jRNsHަޛg\r,v؜.jK3)[EJ2ii{KEiHP^&]Gn8x=K}Wx/KI9-ϵwQ%spܾ[^R}S3$qvq8M[ ozKxcqmJ/ӿ{_}7&ݨ\f6ZSyQz& 7ۉ[8~UNn|nkiTB+4RI8'Nc%tn{!]Ȋo.nEmʱn𵵥J A+wy#+ikǒڂ;՛s85'KmE:Ђu""Iģ5p=БbTY-ͽڔ詻ngL2Q}$de# fs^o{DUUsfwӶ;s1T,ǤtޒQ\෼J=.tKU,7čJ5 N$y3kdSMQU~mO[03 $zAڟsF5^뜞"Կ QHmrR"ӳηer+ҔZ]hE-6Jmt'ޒ=O[sQj)6K}?e4v_KfZheޓ=BV[bY}lݒTTЬ{ȫvO_qpRApVŗ 6ju=*BR)g "O1yhb=tqJ gtm\b3RY+JQ^Ō֍\յ\>+uSi{=x ^w;uӘ#ĸzLn*$anok߷CBӷ}5Yqvdž<( "_OWit5:EZj2 B ρ1̊fi[n!HQF82q1牙nqnEpT(2RMoM4ϳOu ':֧_Xjsg jP^(ڙ{2%E͖j^}ZU[Q$'U) <܂%!s"m R'G5M0<+zM6qYm$ڕ$3ǧH]?o2N<8F1̻r_my[Rf59NjpzBnl7*{.QP 3N&^BLJPjAHCK2Q}$#~YMq8 k(MFMU)8MEqTy+Tʞ-ar5yܕOXw!e;q-Jqܶ䓊Y:LC UE{/t>r"lI9)3KJjϤA 6SEE$d߇3KG*En|P\ԭTn6I-ƍKTj<1H_zwGr19wF N8ݝ+a9ɫM6mhePi%mmD! """"""*1bRKrD"vnrM۫mmĽm]ӡiG~e"˩ lhRTMk^MX["Jݱk7_ޕ*DqĒ&flՒ}`W}~SմZ{ĕ~wm*/{{ѹ_-0ط#P]xlڱ~Tn5wi*lڪ (JxioϏbqKYR|!|KN53 OS222$jzww%i}>N)E+rۥ7c$Ofl/LNث\6H9: FY󡈾I)fB֔JI_ ֣^: 9mY{66㒢7Uj]:.-os[R&gMF3˸#໹kmjq^8W"PΦURjʄWa˧T!͋ lW48JB2ko+ /Nw QwQzQ ے%$ޓ7^YL|r7!v%Trܥ &|M8~ybrn[RV gSn{{*#2#ԽᢏӴHak" ӌcwҜw&RJ07ױ>Ļ =^ BɆ)v32.M1=#6%̠tҤnzqMwԣ~s*%-j|_m*.Yx9Sz=)qE4 3pk+,`=kNRڥ=B=nŔNAx)Q$ԩȧ4z3t#Z2lҮYn$S%y- JzGpu|LBV7ZW#;Wwipܷ%(6jFG5#{$D"uۭ~]֫SrD܃fҎӾ+Tu>-ZTQ& N|$沸ii>eRWݳu'[O̻j8JۻEѩ[]vni= ڒ,[_%kC7I3Nv$4ɎЈeٸoUu:[}Do5|zNq=Tre%ɧ6&~DȍF]ƞG5q m]/w/ \ʲr8=oʔe9U(W"|S]uZd#?Se[W"ֿh][-7Nu:T=)R}.;ml*5Dlf $fF(̏T hiIUU4Szɕ t(%_|2 ~6eM;TƗK[f&]LK^CE2[ȏBOd;Mi|cx,^6;sیGpQ\NuJIFTJ~đArh* B"$H쉩eXPRj?sl"ԥ)su]xpԴY%VESH"ЋJǰ K&5^Ukzׄ8kEgS2h&Se\ Yl]WҶp-ZUvi7QS:4byqOo+[̺腋[6-_Fo.6[7$p&^ _GZԸߍkc.qqoI[9m߸YxOZЦ1uoiSH)P9Uʄjcq= S>֙NeR><;+ڌk%_qT].srNO?s[=vH[]RZHRMtᩗVؾ:/~u)ԍdg%=edVrISb{6vSu=(ܥ)mTv/J}̇8 S3ad:^hBSf؉OɔLhI_1d8,L><_A0y3rXq"'(۱;mFNII.v5_(^q~X>y{3צ I*Vܛv/jW' T'NR'j%ꔩ:mJ3SB}΋!-H-RJBТQoedi9tjENenPpke.%4]#{:>mkEɱdYWl\\\'nRM4&U>?Ќˉk÷!𴪛]]5}UqG~ݏI"O~s6(Ļ)qO~h}uԕd}Q~G,oE!&G&/]_H-O=o{k\̭bkv.Ô܈+;arZx)m?M\3lU$mk-CFXjTv6u' g:Vn_*qk:VC A%'4JV%EY)#BғO4<e׿jQQ]yUr4=wm[K1r׵%Iũ-O}|kC;/VcݩWZ)EHdžTru]8hgĵ-;=>U_ InvTm_jBM+QiF"9*{DI/iuo(=TzϖmPQl_v4z>T*ȴ>YF;ε\t]EH4ꌇ[VrLzef 2T^V>g2~kg5~Nק;{~Z~W}&ŒBӿS2$J?~(Yœ"˲ߩ\O]: J׉ښT{mmIѩn3˧)4LdFZ/zUG>U> n 5& ϴ-KJi2o]uKljvK3$bԔҚV旧iY5.ίfi96v7!v))FJM4{jG~Jt/lUE%pTAFe4qQk\ve۽/u/Im+W')v{\-E|Pms7߮DZRr۞/mu*1ՙaB܆ -xg3#6ۥtRogʌU)׎]ZҞNnŞr}F1Nnޞ;cZ{N}ۿMiuxʉ*3qi'9KHQ$WJxXyرŔe~[v5~/jN9Q4o6rJv FrdxM*iRjMzUinHdн7ᾞS=S'7 } ̽zt7K|_g J=Lq+/Bw_\ۧx\HJUPzQ<hqF[V0x==CsU7q|^ {)Iq38$_A(VgcKu06Ƅ"%i~_ˉk QCܣB8Ku/񋇵u([w}$F|8TՠI.E !;RJ^}MɒD_q2];Ɖ{5}*n7nEInO{Mwv}&q+v [V}Ĝ@%>#dXQ$f;iep.GquixVt x6bj͵mlKقQ[T]zs/&yەnM'W}!Fp_d^Tu N{ɻ'l{խ2.sTu{W^H&;1s)Pӛ6>$mě;Łnj= fLT)>׸+qReɴ[UR\L*P/!$Ӊ3Q 'K=m~6XqW3^W+ųO_[F$rR*u"T%@O +%# ]˽!aܽz{ͷvQh쩎]hGތ5ɇ*DzJDRNLi 4:{~2FmXY-zzĽ^f=]uū{/+&c:Ma{ĝDp2m܍kHș/(--m_vݮK(V{R}.k&yƴ7i^4@3f sK3^Ř˸B=]?gt5KbZB<e;kQLpxuWC}n 5ҴepB##~q= `x]KWF {GfŲ}?G.I9pjWkU]>={7q{kO/^I3==f1ɏ%nnʫ/Zu_yXN<57ۍ'vy/"8넭M2eԷ&Y,в33%IkjMr7xf nmQkX4踼>a-GcIeތw&U=-:qnW)z¥j :WqSZvԒ#j"KrIU)%qrmRoDGQ~SYRsu*V)  ,/x)MFD6O#]z 96[Ui(JRfw'y$GeUީkdMF-ݻ98F2d[o{Rn0n-xsV6Dh|Eb2E:KCOӪv4SJCr"J!!m,hRLD| ZYFm/X~ΧfrN&4Ƒ=Z9Mh.Mܵw/BdrܥniŪ8ɧ|y%œ[M=_tj?F!z5\evM:\ ~F-sg钬OWq“iiȍ<Gi%%n2rqͻllƑ)okw7}\Uk-:&fj솘XerV9yZuʼşdFC=rmo%~ZN78X(N)_7.Εn1MpJ}62jjJdI";R5&iLԸc:jmqiQj$ujp\{;v5B񥍪Xn Ą4qOERjzN(Ga٠䌡)p*v(J7#ZۻZ8O W uONb+^Qipv9GvֽƼϯrYƖKGJQDNPhRJjᡧC"21"9ѓS1;R_O7/WGz)8fE%F2ukmvSov/iZ&/]~KmI[:^~ͤ\kMi稜\ywJt3W7 8Ʒ~ݥeFgѼw"8VVSج\뻆}ݭ/J6Q)d|)zU3>k\L=;ow֯gN3pKѫ|wmkZ$z^2R:E)f>ς нd|#׆?\ǔpV{;\$ƵE%-ͪm0S6[n< kE[}mvE4DDZ^$OZ0*$~XUv҅B@^?]so#%ojw;Y#SxxueBگy v^i-)s)zV jC{7Gt.w3v,ygg8s]aE_,*E tY5k٨h=o"m泏:\6w噓aiL׎n^c\75AGkЯ0Lf46َ`egZ˓p/k;̛]kq!ݸzpԭG"}R9Ve>ˏHUjJ-&7nrnwG*Xv\˱/vN}O)ʼn&CV͍f̵]r\PMB-6Du-#RͰtRN^)mT _}nSȕC*_xBuTkJW[`ɩ`ejvsngP ڻ.-WUtܑqԹQj)t;vN&RNũT+8%IXӃ5fK՛-d9 ]CƑm|nZ-6=Hz,*aEm W3VzRšdY~Xf׀Xx"]s;)5u*ُHB BRGS6bݶؿ 9j[1*jױga7oX CUI%0v#~\-O-Ꙛuɷ쏪&5mY٦M`LJ2qK~HZbr =N'YobI. (^ ׾{_ ?OJ`S`3BN[}5w6:ǵ/iSlt=4F*d&T4y/#. ɵim5Uֲf 眕6Y7 fơ=3dϕq뚩$qTM-%r!$@A? ޾V0c~{[{;򥧅a~ڵ»&ڄv1ek=wb MLkNAԬw-x>~/r=e73VeVN)K%Sښe"+3uXuچrn ֺVzscJ峻m}vb㶓n\YbIUBT%*,0nov=;z꣓S/nSXSpl##k9mXGrZv^Gde!ŷRԠzQyjC]`gToPov{j~KRBMY}i[߶9KL2ԉO0K#m>wB[ٍ+n[[b٦DX ݲpo] [\m5qdT()mo4Oy9Ie b][wղmM~vmi۱~t \}$яimRk(L c Cvk7r9_r1 ;zv|F@KyZ[&jEji/"6$69ml#e]9s\{ScL}Ȣؿ0q/nZ*t,CLoD߉Njǚy=Pgmu6^]l-["çUʖMlʍp-"qmU>۷uFOJ%Ǔkx 'g=睋k[3u,{³WɘݪF]ՍeFX"Oy\,cچ=w/gn Ļ]#2? vqy-gXnR.^}ݺFs{ŝG]}e|#0mjx"ƬWكm?rgU^xVB":Dt>@LRbun~ݭ,w+v⪕;\U(RYa61>#Jm˞Μ9g9XKaG='u8gf}'qy#ɉw J]We.ʲ-<+&q%s?2dњztҼn`cΤmmqMdz O[-ߩӲ&;[tmܝVnr">{x<8U+p:Ig]zjGkt,uzf}dؠoJaکqEq -(:d<պ=eKy[˗^%ZXkX[C2߱\ITTLGzANM￵i]K>UsOGDDD.ZF6* ҃V Zhz{'xp^`wo8r0h ZmJ5"jb[l=yUu7-;7IT%:jFjߖm0tzU'K)څNۧYJ)4IQ}^KWm7kSP>q;ނ#)'n7&׊r?óM{IwR\j2Qn[v pe#/tAF\ϵ225q֒om6z})6҅*oqDsMf CNIN=T S2t,_ѧ}kveMF0J\Rnnݙܹy[rUc-j{yGtkQ%s]5qB.Nw.JN1LvR Ui5J ZESQԙr):MJ+g}χ!2;q([jAud][ljVK3$ײSJI=/|&tl'*n۽f.frܥ jQO8>&Z];.|7T/C}$ڋUmP2Reҭ8hFF\L 3~e v\۫]ݝNmrnB%*]Z«hKc=BTLG :V74$=Ǘy+EX'4tn(I:Ѝ;Df8c,k1%dJ6.j6ź{N~l6&*fœI7 WAlGOu-ҢH,,(ǔe뿋쩨kM܍ZſgRvQ' 9)?n|er˭|I|-fGK.rΛp8XV1%K6mvG+tc+qE&ǸC_Nm:l=_/m5^[dߌڇ.c<%:)tQ$Ow~-aY;UJ>=F)2[nk؆?훐M=l6[4(O.]2#-H^n#->&mp5~Fӛ+|| S,xag%qkEUzUgæBhߕP(7]kFnq?֖CpruZ6*rEڊtS|*tI*E}7R<,nUU֫^I7Q*mSly%rdȓd8hE<9oHhMfNSRj[i7D[Rj݊+kდq{"$$H?p\̅S?㭻;t~R߁)^/>Qj`yt[w ԛ;²~+ߔ_ YW~|o]?x^ᯛ `ʼn;g)T@vWn]>&4lp+$D̢1l|ȨF%-}.9[}w~ ԠLM9hСablfe&QoW!s?wjLK?s7yO>(=C~_nyǜu?v3vyo oI@qV-jeES^[9WoSܝh"l2C1a͔CiJ@3:Pճw=/7ovuk+\V;lDgն<[A+rX~d;m!_s8ݖ׷;;.0llUC+?i#_crʙ1~C.\–q ul8Hܶ2m`ܻM3Tov|Bs rɵ"oLS- DКw=Tv@f'6|YlD͓Y%׵-#Ѯo%:&!3o%\J<02;K87>^vgƓ# ;ݝmz^Y6=PS39U%~ &f# }o!muH;ʲŇ˷yvP+&.7e[3'vR4Yj̗IZ`e˽3o[WU{ m[sUbۋZǾۆl6~9'V*.\S2<Sd*zY[aŶ`]C$n.v^Ʌ dng>ەZ,Mmϑ :n6nϦezWqUJ4! ۇ4R! =>>Fn|Q[{pRO17ƕ~._I''00k=b՛o}Osðc2'o\3}ݭQ^2 . R1yKȣtAݿ-uܾw!`?1Whn|gzUo[ECWwjUIן)^h#1ɭ!/Z np;o;ΗŻkXs."6E`Z1 עӐ9Kl8qd q} 2Stt;#j>;խabONŗ=fwP1j)l6J̶|gV2`y/0E˛6+ԫ1? 6}KW c\KoKͨ2ۅFw–s*TԞLיuDx .kCzWXhy۶gLu|%TnupǺl-S* PRaLnT+c+*xl.v!.U=|; !_L̎뱚U=4hm:ٯ"y)$:>%(n}X'p[ȴ ^˒4kƓmzDx \ 'NqamP7nyN݅=j7%McSڵj%STy qXymvCg{w/w=wSW5r̹u erծˊsOm=DhEҚRb#n)QOxtվQwe]I}wCa'"[ۂ-z}2UuKP$㜉ԧ:mc<Ý>RoL?wu|%ҷ&K y_!y9 ??:tq3(UU-lkS'ɸ@jdzQˬR] EVPW1DJq2n:,c|ǻ̑;y{X,ۂ.u.b˕u.tKBjQ"[S園S`ٮdNبeJ&9Ơ ~0a(Vm٘L+Jr*vڑE( x0+tp˕ n';wm-ޜMOxX>{#2%jgb2M[`K*\5@8l'e=0u+w ֘鳾{y܀:R*Ya]"Ӧ%ktynlۣ65,3gU}{GYrb;ge'TKwǘ.,rpܚV]Tr,!dp /ԺU,xՉ>s׽~W5oTh yx?xrrx?)?ilbT׬,z$Ԏ.UH٠\U1pU:]JwSrGZq8àd驐,N67QYBӢD㏙W!Q25ϸo9ms-7-%3CihO.J鯽-;MZM8ku-7k9S$8]q2E(}bۏI[DKOK}3KUB^u %Y,u.-&f#]'܆o$x`Yu,dzwM;#oKxn;\[d7}Rb+*Y䛂ZuBӱl{j0O̓}LhK;[aֶaGL{Cb#S.T[>߃F]NK"u^LUʐ_ykW?!GRj29͖qa'0[npcDvV)qz9R)PۨM^aJx W] r>];eN3vxdmĘ(5W2K1䪖weF{mE/QP6\u54x5[hۮ-Nk”i[lUgL]J}5 S:EhiUrgHl!ŒJ$pe=q^b͵Q' ?6|R\,JA ڵ"TDꈭ:ymg`B5t%M] <N_zv2_Ortٵ/i/ReӮ*7[qүqEG* m"[I:6e^p"I$jԴęh!m)]GZkcjS!{e^z}+Cѥ9;R|/ֱeiUԏCNu2Zhcٗg$ݭwvr P8*7/Lk~I'Km1+MW%Bk|oOm>-#qj*|Dbѱkn|n{v#jĮqNpMIUm(7Liz;{ҜݞڝVƚVϬ+sO!OstGvxӉ']uӎ4g_ 1^-8ۦ k!)Ύ5O;YSB#2Zzχ;<.ֵOtge~.(RC#wFZeGZٸ6FFJ4e2ˇpJT$[wgV)q6muDGJ56q\I!̗ y/I~RtJ9kJ]Iy*'FN0s.[l!fw'y(7$œ WƫgyΙdMEU JQJv̋vmrۖ.jWR_M֨djYgSj0^\y'EoECjm$ IƩK>Z28J2TiJ2N#}.s cArl嫶nB.FIJ.)۔\ZiM>/hLĸ=C1s[?YMqp|94- 鮝𦔽/k^#NT(Y LS$6˩}{;5 )B۷W$qpN)qqoot}ZDVә;7TiK|6f3h$dԄ}fqݡ>Nb򗉉+ͶO]>ߡ_VtYf79ڰիF sq~prս|QM)g%l0ocJȨHz V;Bb/kLAcfPJ,ԭ{ƍgpjNR6VSI*$!yV足jᇑ.](EܣqM\qJ2eZT).<9UB/(B0j)mtKEj#׿fDI-=rZړj|'Nڤ]k*i$5qt"ݙPM6E4ke^Z8ۏhz$Q(R Ay2zfRñnpnkbkI:=j &ΝșW?׵d{+ύM'??XqeeĽ.[o=UxFS=ӷdZwenՄ]_X=ĭVa* pKs0ބۍfJ3 gz̚i|wnxtjc¼5${(1fXQ65ȼb̶Zkn>%FQMJXӡ{TZEVNᖣimT/37cNJUPnP҂ZOE~"-Rc4^b- FEͧtf5[)S!OZIښݲ͑;tvܡ+N)AR=hCNn;wL16-:特7M$=Tҕ-.R[HٷnXk sn[ҞD-0WS9p9:-Ϸ-jѬNu{ҹfv)[Ľvwfg(ٷfe+0mYj8Q1\ݧg]Eǎvڿc!4#j5̋C2"}BRriFp7=ô\TZ:\BLfj#I22װ<;صZl j 6:l"6]۸ K'6RTѯ^ئOԓV\?$x7s#r:Oh{ց=MmuHԷd{pN /܅:UE#Yy+(SgQ(Щ)RHzw>^Ѿݻ>mK&^ '$Jۻ&w%F|xfz%˳ L~3N?Cy9 v w/{ƿ kz3x> sXv}vP"@WyC z`'톽Dw%-tt yVY\wmuPYQA0iG-2JP,6/gˢ]u.-n!Zw.N7Q]Df}Q0({a\@=i_X7gFǘ8^⻲}G MZ1)WEfO12G+=-B@z\`||w6ċj߬m}UwRox֢I &c~XGP6Qndpvܻul'V7^FJt^{b^B(L~sѣ6@߿^xqU!ڙ5|Vpvef-uӥ^3  FSDɯKD%0r}FF穛r7 +o"V8tv̖NQU!5uFd"bCr^bJ=֤fM#ʳԷP0O-9xRBm\=`r-:;~3Tl(nXtXi%2Vٛ#vwqƴ`L@"H‹qW.j,JM5B[)WܺUeZFqc'V˷1W7V̾-MHФwn8N;HPSdݷC7&2j.W\τGŎ'Vb]c.x+Rx1%C2T{myg[qU|+m:M:շ8҉yWd)ՋWS%%:iqlʹmGwݹ WnNŤѩ5(9hTٵDdGUi-)vSs2 2{OnT$Xck n:¶(lASLeȔBjμPpTb2~N2~%^k[ܗ[Jzs0ӓHBKq[}JَA-$dFQgjxxFv4r/x*Rm% `4J(&iv7SkԲmSH1YWmx 8n.k']:Z˭_W >ڃXЩ. jTq%Aā[E}amc]D:rmHRiu:uӚӢ\p(5-q%e)(۬ҖȽIf<߽pr&ݫVfY91q2ĭEQgYbTGQ&,yL+N$[q*RVۉQ=FuTܻ>f>f㋳8N6$܌n)9&»iˤsX,݅܍ȩv+sRTpO}d?Wn/Inpȸ%O]StQO|v5\}7Zwb.AIVK^:wb{[uݯcytO߶S<{8KSRׁH̏N7ۚ[xkwYy_'ZӵF+>쌛ZUĦreE9F[24De{}@:ExWs-\ǻ7K-\JNvEk%:s˙#κ].oͳ;լ7wB6nwu:$L; DkI#Wz.:Xp(˅v$Sq,wn\qIN-e<5Oe+vuYTpcojUI_ާP8 O 7&VL8z$_B-H-[uh]T{|8=qVRN-:Ij:7PUtXϷmy鉿:RIM~33ӸS2#׳GdŲ5+/Bx{(WzȨ5Y㞎#|˖+ ط.|e<o/rߔX>7s}VE.OVti׽ .5nNJO"95{#q}Ay9do]R"M6z\tnNS-D!@3N_jicWsy*5uٮRcWv/.,j}=S)j5C^> Ie =gu9ӛqjtz]۪TMoߧI!Ǧ¶m:,"[L!{qAv-o 3{"KʼnrIkfٶj2ƙ؄S`7` k6jzޞ?e5G&6uʷ2%ԒRKE*G\Npom F/V |C0.q_eenƣ<5Oh'67ɪn[SĽ{ڔjǘzs;~׌(ۂ`ܢ1ƣ` _l9Va6%UQWh~P~\F^ZHR@:ۧCJ{ôGeBh;~ۧnU J\O+n2 RҠ)ng}Kh{5+S×ܛ.1ZjG)iRȤIN 4%{oΜ/eO[Nffd ĹK?nnԼMqX'܌nZvq<ķbFnͪaQ`5 s,M_լ?-@_{w{ӺձJ}GF[%v\5[ŒGkOw/ΜM9rjË%2+rd~+󲕛C9U۳r[aJǭm|˒LAʨSCq[XMۺoubfp:t+ΤĻo ][ zt-*67kvS7D·MMCQXm;)܎n_h%]4ܙnRk!]ڵsDUF"`R, &#R_*[z*ZqFXɻ]7|۵w+'pFDەs=r./ᐚm3Hשy yD"jHCr':sA65نѮ^o1V/ f;nFr3VM)e*- s D'H݅fӧ\*޷[k<7u<-]֍Q8R h|p=WlW3s%Q %3l}@U-K6f-NϿu|ڴmWN׮[׸F*mW\%r! C78:޳vBG7ŵ.JթԚ2x)ST!řn~9 W:Wpܢ件{xf8ٳwKE ҰWxVB\qBZ 2wMb[lGSnyԚ~z9ZmያvoN2Afnݽjf>)j3 !;gOYʹK" Wftڎ+׭b*2ϻK>ۢӱeyԪXISUm[z+ugX%0lϏnvg!;t{BqPj>PyvR7Cj]O%+ݲ :qiMj6W}3vC/R=4Som]ŗ=ю, TF6U_-\6MyskwMr&Q\wjKܩyMϣUj0*}RZܷSdY3>Zjqj6TgzpA/M`/Cmл,޻feE[/+uk^Vs1W$G(JsW2ٰu*߻q*Y޵.Wi:ur5T),=0uRmho.twܖiYwrWHntvEj8qhf`Ͻpf(R&>Ki%I7$QӖm-2 ~yߗQ-앑/ x[k8nw.c㩵k}]FkbJl:{.(˩n0Hqvαp7 귎.Gupx[N`Yq'+ruU7[ү+>!xrȫoSo]OC# d^Q]\>!ƛGw^Mx"-+%vdX-:M2UR%d>%l ioSu6lsj7D P>XxHz Ukà(n^Q V>5cVtWj SEiJdznyej[lE' 3kuٌNn4JW)gB {4 j6&]' m-(ZMEz8cz>WZ6#7+[,MR-Z!4ܓtCyE|umj1ƽvƷV\;%>Q :#Le(iVz5 4ũۤUWxX ^(ҔsլB2w-V ^R+; ˂M\z+Uwr+RWY⺧~ Q*JcYSNSλUd8in=v K낫k\IRרSUaCFmϿ5̗P|u ZTԕ}>oYѲ1sfP+sQkX8Gb~6r,s>^\,mGL+7[n-E\.Fqḕcl*Jmjb5 ,m]c}NXfeVlǸJ5eˡ$4%g~N p4Y*WwW٧<8v#;qԩTut,m"#Y D\5V`\\Lȋ];LȇiS6ϝZ l>LruR\v=ǘϔDg=ԈdFZ+M{=|,[;0>RiSi4,S5}yxw&(E7&fݙ4UՕ! ~'Id)]ǽu2K-fޭ \08Vڅ쓬=Vy^^ IhyKR-B#Ըr=]mܻӾ'*Umkoy rTqT_i,/8Q^<ݤ|4ԻO(܄"'5N~#m.(Ҿ2i6Uev&I*<}҄$eNtÛzyWJubW^iBW.܅Wڮg]irO6Ve90sgv.+sV޿aޔ[p?3q*FutUo*eL\KM'EG*ZcAFfG5J 5jj=MJ3OK:k˝'NMB7m3uFҕ\-Ywg%PRqMIyZGY9|μvn߻5cWݷa^+X֥vnݘ\v7m>Fgzv"-;Ew֝}1|RjN𿊀7g#֟*GQQ|#/bo]p$>_Un9гUbn9׃ErQBU-^vDmVh'<R[fdHT]*~}3j;nvjc7s-rӳ Y8[n[1pJx kX[Jk9Mn!_Nю6x:iZ˦U |߉^Ԛ݃hYxk &U^bwKk.[jE+P(˞=9j@snCv7%c_7=xǁ<l {t'酚+1F‹l׭:ݻILruǶkL-L(K0L1&>wXB(pm;1fpnlp֓%Skidkt(U +xulo'/ڕeN r=^pZZ:Pnj8Hf"48ijY[ N[yZٻ+=  ø:3 ?^ܷ^Sr#YK[UF?CuhC b]GM')mڏsNrܗI]ljq6VB. W,UK"YX5{c >Iqā> T:n!,5l2VzCl|+I[*SrjnS6٨y+x,@>П.g+!rn9>N|W>OZT_ut Y""v7|sfި;Pclm EùN,{'fNT%U&LfH8~1v>Il}统u6P˗c(WV~H^bMU.o*oOF0N:_:6Smr_.b+|ݶYY غF,mwjv>f*>QM뭱Sd:`N{l/⎱;n-z~"Gze퇎J5S KG9!Gn;N1 ݎ h6m|S?ɂ5'WOÞ 7|7^ao @mxGmi^jϽ>01Mf0լD3-2T. VXR"ɥV Kl J O7|u?bvа;6.eߓ|[1bmRr,eRz`z 6܎-ͨku͹Fː dPhYgZUj}nvX;z=gVեTv_J }\1n7w2J?ޘγc\E 1Aޑzq;\r]]\Y&[nsNei\uURje*Qk2CSl*xJz-xٶlm+|UjUؓ`Ladqiĩ!Gd\W~fz;Tn*PdRM&T4`չSWq5k훶(N"Ӎ% V]֦wb.nUO!u*J&Oӕ2e|Z=eV쫚΅g#+/RW:طnbi*Wyo)p{:ETKؚR(RY+r웓r(IF) VmȵNB:h Q1ғ|u8E]{,'$-TR[j49l*3"I鯴zhd>Q+\BkNF=.$ZR4Nwհ(IpNi.(Gi33#33e$FXK*NdWrud[r{xnk$v2ıh+J1TQ[#JQl[tRO]LHKٮ NӍnF񨔤֞Em'MILB"ԋ%dBŋ+p̿_17jzT~4pc Vo\ƹb9Rq-'1j;8ܗ)hE%DZKS<璸Bu*%*Yw5ڻ9ۣ^z4U; Ñk\U(o~G?VUĎ:?P?_F_Kߤ~ᓾI |pr.Ok\SklRhҪz{­P .}SktZ7UQ4ڌIM8̈eaӊJZ%FFZu,KZvln廐SNFIVtuNi?CM5]+Ph,{jN JSR$IS^tSUVrORYu.9WyP6 [Kiu m!X|]Y79ӄ)\ģ)pbڳr%*&ꑶ_-H*dzk)1 V3')UAϹٶWRxe'պn۫h7AR9 EAJeGLms!%D| A 5]/Q3eb̄vnVn%za\m kZnv([emqrIҕij|""><hjJשvvǕ|Pޟs}V~2&Z?+2N&Z4w@)4iSڪ_>/JN9Hiۏuf8'It[ȲR.hZ$ȋ_Y ~U<UUO*6b)Ovzڜj\R̋.$FsQuҊj^נ䈈y<zZIuP[}Qm=C?zN(Exqu/kn S-FzKZzOסӽjJ\)F3b!r5ٝ|;6 o=-3*λ]αb\abqRi-w޵⦪~b8Kpo)Z=>)ғ"5/GTZLE-輵f7ݘ۹~+&+w/7GFI:l33fg.N~۲\2|*cnermnnM+Fq"ѪIz%j =YW8@~gc/~?N'?)«qȸs➟n=k" X“m֮VreMh2[uݖ] *FܖN)MȐ`f0 g,C9̑o;ddudJ=In13:ݒvvdMUEJLp^,6t-@͐9'{7m{-3,>hnF;ѰM)->>+Ěz!R* :`e--m7nB\u{b U>[8֪]6^ߤLʦ\DFNo$$dͶlgno8OrsQ\l̯hRo8tuNo+ CTxu!2[>ctFpeޓƻֶR"3QrQuOѳgwQr;S~)6HhZw/GgVTmUf_yt7%$];zLWF̰xy2Ʉu!MCmš_0[W6jf#a-KLi+3Q7c^qg%s<1aYIQeZf+}>;S6L0]Yu_h9߻<ƅpmiM$AVvŚ,*#t2.8Y)-Zhshü97/#Oro"u^/uFgWɺ,p:6a,^x%$Yve^3PƗMnTP&yS}OJ '덫MH^:rXԴJۋ/rI;S*,+yz1hv)Qw^ڍJ2oL׊q(\fDj:^T%vOadɂnS}ZO)N*λdaȜkG_PIEO}нa(^iQX᯦-7^)%g'SJx(.S9zVɴZ{E ))ۅi/s7 VIV-|sj0*UBTHIqRf>FP$KqN0 R̻8j\GcC}IUz\i 6F)Q{Gҧ3qSzKj-Az VЛS-zy:8*mNk|D鿓ND2u+0Yŝ7kqm·?8Ib]u>˗^_>(]vӋzv+ݩ){vZrJ2RQ몋C$z [,pp,8mڊbR]Il .f~d/ݓs㓓mͶ{mgjQwn=Oic9ܚm4Q/6ݨ[TƧ?nԶoytf{@AzT{e{[O'ZRZt~AGD?s3􌿂ՉIw'|~U\ w~di:Kޱ)U/sU%njѩ&GSP^ǝd)..!^U` 1wX[aԇSxoFV6_扐)T 2Mfd=ۖͭiZ7KK Bi9%7@<3<ճԻU,},a}FRqɛr i@ONJvK KLN M, ʖv0n-]DwlI-X6ܶ$Jʴh5O+mOI+Ra瞠\ MG7BفjYo1#͖0V`Ѱ2M?c8>-Crt*JkIGS:e#hPKx[鱼>{5m;wcն&>j-M֥^َ) 6yȜl_w{-ō̱r> U=]iw3)r*]:K]6BdCTZ|>gf}LW}[$'Y5 &c -j.z6R 67MԷFMnÌwI7w5E}o޽+K ֵy4܌ȥW"COyR[q5Ӱ͙f[v"_#q{MV6܍3"u9BK(41ӯqˇc${ߝCi6I(OmθzҜ5k^:>Jzw.>qV8{vU[ڶEm|DžBz].KHjI]x;Mɗ{m,qZXr忇2u^RO2Z}ZێS[2Jen!*NDcrBUً4<ǼMҲs1Zw57c3&ĖڻzmP*FuJG1-dN:|OU}ҵgi2t~F^^Z.VxjvŧnNNh<:]^~NN+ge^g.SԔGFe߯'[vn'(ScJ]kܗ7eJOlRrfziݮq̋S"\*U<*W]k$FջV}? 7g#֟*GQQ|#/bo]p$>_Un9;l S VvQU%OLU{οmU6bZ1MTx%!֙Q7, J=!3 ;Q,ڌ;6ͱ݅q^&ߔ·n #WbwӖX.HtG)N&d̵zpI,n cu ޖUj+VXUp[w]N o.J6Z8Ts&utxln;~HPHS/xw`G\ʡ¿rj Z^vt"[L:SD\h0sUwR,}[x^X,R2Vn< ]2YDr[SRKs8tXb̷G?Ps Tv 3be,zVz D[/I.KOEQrm'$7|[J>r S`5յwT#\w1FTz\Ԛ &"ׅhSHrD\'r]~/>p;:Piuu:"9ő=tTaS7V2rӷk7mb[^WmPp*[y.Þ6f]cizJCgRR@UVl큝.WJP1N{/\whZ ػϧӱE7|E֫Sί.x-Y&pi%v''-x6r'Ws*6=DwwUu]=C?MK [yrtܒG$!WGqJ*%SAz ED[^)/tė/g=#Omd.|^n/sl׉g DZqemqowݮRzUܜ=ڽ-o/Iۖ;qVʘgPp|mm;6zGl9.8pwWgsJ2qPbe}}UpNjٯ}7TMQKrؽtEx%v w߾8%|j;~|}pK]ơ/ w߾8%|j;~|}pK]ơ/ w&~e_H 8PL7:%ʭ5Kw&U2vwR_+rm'}C7#rWoO&HoG?M$UR7{FU]u ;# !Wk`|W>׹潇9Vn)6)*ҹ{%qV4q>W1vi#T"Qk&GwxcJBJ- Ϸ^ˁxkU}ԣ/3.;]J=<*)cS)ROK9H=,r zX @)cS)Da^ԽQ gxJI=w֣gf*TRj

(E-mail: jhs77@korea.kr, Tel: +82-63-238-4657, Fax: +82-63-238-4654)
• Received: January 1, 2018   • Accepted: January 16, 2018

Copyright: © 2018 by the Korean Society of Breeding Science

This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

  • 5 Views
  • 0 Download
prev next

Full Article

Download PDF Download PDF
  • Gayabyeo, a Tongil-type rice variety, has been known to be resistant to the brown planthopper (BPH) in Korea. For genetic analysis of BPH resistance of Gayabyeo, we developed an F2 and F3 population derived from a cross between Gayabyeo and Taebaegbyeo which is a Tongil-type BPH susceptible rice variety. Based on the previously detected 284,501 putative SNPs between Gayabyeo and Taebaegbyeo, 99 cleaved amplified polymorphic sequences (CAPS) markers were developed, and they have been used for genotyping 180 F2 plants. By comparison of resequencing data of Gayabyeo and the sequences of already reported BPH resistance genes (Bph3, BPH9, Bph14, BPH18, BPH26), it was revealed that Gayabyeo has Bph3 and BPH26 resistance genes. Two InDel markers, Bph3IND and BPH26IND, were developed, which can be used as selection markers in breeding program aiming at introducing BPH resistance genes of Gayabyeo into Korean high quality japonica rice varieties. In addition, BPH bioassay was performed with 180 F3 lines for BPH resistance QTL analysis. Two major QTLs were found on chromosome 4 and 12. The regions of these two QTLs included Bph3 and BPH26, which also supported that Gayabyeo has Bph3 and BPH26 resistance genes. These results would be useful in accelerating development of various BPH-resistant high quality japonica rice varieties in Korea.
  • Keywords: Rice; brown planthopper (BPH); resistance; selection marker
벼멸구(rice brown planthopper, Nilaparvata lugens Stål)는 주둥이의 구침(stylets)을 이용해 주로 벼(Oryza sativa L.)의 체관부 양액을 빨아먹는 노린재목(Hemiptera) 멸구과(Delphacidae)에 속하는 벼의 주요 해충이다(Seo et al. 2009, Sogawa 1982). 벼멸구는 우리나라에서 월동을 하지 못하고 매년 장시형 성충이 주로 6월 중순과 7월 하순 사이에 중국 남부로부터 기류를 타고 약 1,000 km 이상의 장거리 비행을 통해 날아와 벼에 피해를 주고 있다(Otuka 2009, Uhm et al. 1988). 벼의 아랫부분에 밀집 서식하면서 직접 섭식을 통해 심할 경우 벼 식물체를 고사시키는 “집중고사현상(hopperburn)”을 유발하기도 하며 rice ragged stunt virus (RRSV)와 rice grassy stunt virus (RGSV) 등의 바이러스 병을 매개하여 큰 피해가 발생한다(Hibino 1996, Wilson & Claridge 1991). 최근 20년동안 우리나라의 벼멸구 발생면적(ha)은 1998년(164,577), 2013년(104,599), 2005년(90,500), 그리고 2006년(62,279) 순으로 가장 넓었으며 2005년에는 집중고사 피해가 발생한 바 있다(http://ncpms.rda.go.kr). 벼멸구는 우리나라뿐만 아니라 아시아 지역 대부분의 나라들에서 심각한 피해를 끼치고 있다. 중국과 베트남에서는 1990년대 이후 벼멸구 피해가 줄다가 2006년과 2007년에 다시 큰 피해가 발생한 바 있다(Cantindig et al. 2009). 중국에서는 2005년에서 2007년에 걸쳐 약 25백만 ha의 지역에서 발생했으며, 베트남에서는 2002년에서 2009년 사이 매년 약 400,000 ton의 수확량 손실이 있었고, 태국에서는 2009년과 2011년 사이에 3백만 ha 이상에서 발생하여 1,100,000 ton의 수확량 손실이 발생했다(Fujita et al. 2013).
벼멸구의 종합적 방제에 있어서 저항성 품종의 재배는 중요한 한 축이다. 살충제의 지나친 사용은 벼멸구 천적까지 사멸시켜서 오히려 벼멸구의 2차 대발생을 초래하기도 한다. 벼멸구에 대한 저항성 품종 육성 및 유전연구는 초기에 국제벼연구소(IRRI)를 중심으로 수행되었는데 1967년 벼멸구저항성 인디카 재래종인 Mudgo가 발견되고 1973년 저항성 품종인 IR26이 육성 보급된 이후 다수의 저항성 품종들이 개발되었다(Kim et al. 1991). 우리나라에서는 1975년 이후 밀양30호를 시작으로 다수의 통일형 저항성 품종들이 육성되었으나(Kim et al. 1991), 자포니카형 저항성 품종은 화청벼, 친농벼, 친들벼, 안미벼 등으로 매우 부족한 실정이다.
현재까지 벼멸구 저항성 인디카 재래종 품종인 Mudgo, Swarnalata, Babawee, Rathu Heenati 등과 야생벼(Oryza officinalis, Oryza nivara, Oryza minuta, Oryza australiensis 등)으로부터 약 29개의 저항성 유전자좌들이 벼 3, 4, 6, 7, 11, 12번 염색체들에서 보고되었다(Hu et al. 2016). 이들 중 Bph14, Bph3, BPH26, BPH18, BPH9 유전자들이 유전지도기반 유전자분리(map-based gene cloning) 연구를 통해 분리되었다. Bph14는 3번 염색체 장완에 위치하는 야생벼 Oryza officinalis 유래 유전자로 NBS-LRR 단백질을 코딩하고 있다(Du et al. 2009). Bph3는 인디카 재래종 Rathu Heenati로부터 유래한 광범위 내구저항성(broad-spectrum and durable resistance)을 보이는 유전자로 4번 염색체 단완에 위치하는 세 개의 lectin receptor kinase 유전자들(OsLecRK1, OsLecRK2, OsLecRK3)로 이루어져 있다(Liu et al. 2014). BPH26 유전자는 인디카 품종인 ADR52에서 분리되었으며 12번 염색체 장완에 위치하고 NBS-LRR 단백질을 코딩하고 있는데 BPH2와 동일한 유전자로 밝혀졌다(Tamura et al. 2014). BPH18 유전자는 야생벼 Oryza australiensis의 introgression line인 인디카 계통 IR65482-7-216-1-2에서 분리되었으며 12번 염색체 장완에 위치하고 NBS-LRR 단백질을 코딩하고 있다(Ji et al. 2016). BPH9는 인디카 저항성 재래종인 Pokkali로부터 분리되었으며 12번 염색체 장완에 위치하고 NBS-LRR 단백질을 코딩하고 있다(Zhao et al. 2016). 또한 12번 염색체 장완에 위치한 것으로 보고된 8개의 저항성 유전자좌들이 BPH9 유전자의 allele들인 것으로 밝혀졌으며 4종의 저항성 유전자 allele type(BPH1/9-1, BPH1/9-2, BPH1/9-7, BPH1/9-9)으로 분류되었는데, BPH1/9-1 allele 그룹에는 BPH1, BPH10, BPH18, BPH21 유전자들이 속하고, BPH1/9-2 allele 그룹에는 BPH2와 BPH26 유전자들이 속하고, BPH1/9-7 allele 그룹에는 BPH7 유전자가 속하고, BPH1/9-9 allele 그룹에는 BPH9 유전자가 속하였다(Zhao et al. 2016).
기존에 육성된 벼멸구 저항성 품종들 가운데 한 개의 저항성 유전자가 도입된 품종들은 보급 후 일정기간이 지난 후 저항성이 무너지는 경향을 보였다(Hu et al. 2016). 그러나, 두 개 이상의 저항성 유전자를 집적한 계통들은 향상된 광범위 저항성을 보였다는 연구결과들이 있다(Hu et al. 2013, Myint et al. 2012, Qiu et al. 2012). 그러므로, 기존에 국내 자포니카 벼 품종에 도입되지 않았던 저항성 유전자들을 도입하여 저항성 자원을 다양화하고, 몇 개의 저항성 유전자들을 집적하여 벼멸구에 대한 내구저항성을 가진 벼 품종을 육성하는 것이 필요하다.
1982년 육성된 통일형 품종인 가야벼는 벼멸구 생태형(BPH biotype) 1, 2, 3에 대하여 모두 안정적인 저항성을 보였다(Kim et al. 1989, Lee et al. 1986, Lee et al. 1985, Park & Song 1988). 그리고, 우리나라에서 1980년대 수집한 벼멸구 집단과 2005~2007년도에 수집한 국내 벼멸구 집단들을 대상으로 실험한 결과, 2005~2007년도에 수집된 벼멸구 집단이 Bph1 유전자를 가진 청청벼의 저항성을 어느 정도 극복하였던 반면, 가야벼는 실험된 모든 벼멸구 집단들에 대하여 광범위 저항성을 보였다(Seo et al. 2009). 그러나, 가야벼의 저항성 유전자가 무엇인지는 아직 밝혀지지 않았다.
가야벼의 벼멸구 저항성 유전자를 탐색하기 위한 선행연구로서 가야벼와 벼멸구 감수성 통일형 품종인 태백벼를 resequencing하여 두 품종간의 잠재적 SNP 약 28만개를 탐색한 바 있다(Ji et al. 2016). 이 연구에서는 발굴된 SNP들 가운데 제한효소인식부위에 위치한 SNP들을 대상으로 새로운 CAPS(Cleaved Amplified Polymorphic Sequences) 마커 99종을 개발하였으며, 태백벼/가야벼 F2/F3 집단을 이용하여 이들 마커들의 유전지도를 작성하고, 벼멸구 저항성 QTL 분석을 수행하였다. 그리고, 가야벼의 resequencing 데이터와 기존에 밝혀진 벼멸구 저항성 유전자들의 염기서열을 비교하여, 가야벼가 Bph3와 BPH26 유전자들을 가지고 있음을 발견하였다. 또한, 가야벼를 이용한 벼멸구저항성 자포니카형 고품질벼 육종프로그램에 활용될 수 있는 가야벼의 Bph3와 BPH26 유전자 선발마커로 삽입/결실(InDel) 마커인 Bph3IND와 BPH26IND를 개발하였다.
가야벼 벼멸구 저항성 유전분석 집단의 육성
2014년 하계에 벼멸구 저항성 품종인 가야벼를 부본으로 하고 감수성 품종인 태백벼를 모본으로하여 인공교배를 수행하였고, 2014년~2015년 동계에 F1 식물체들을 온실에서 1/5000 와그너포트를 이용해서 재배하였다. F2 집단은 2015년 하계에 국립농업과학원 농업생명자원부 포장에서 30 cm × 15 cm 재식밀도로 1주 1본식으로 심어 재배하였다. 생육성기의 잎을 채취하여 Inclone plant genomic DNA prep kit를 이용하여 DNA를 추출하였다.
모본간 다형성 마커 개발과 유전지도 작성
선행연구에서 가야벼와 태백벼 유전체 서열 데이터를 이용하여 두 품종간 잠재적 SNP 약 28만개를 탐색한 바 있다(Ji et al. 2016). CAPS 마커를 디자인하기 위해서 자체 작성한 Python 프로그램을 이용하여 이들 SNP 가운데, 비교적 흔히 사용되는 제한효소 17종(BamHI, BglII, EcoRI, EcoRV, HindIII, KpnI, Mlu I, NruI, PshBI, PvuII, SacI, SacII, SalI, ScaI, StuI, XbaI, XhoI)의 인식부위에 위치한 SNP들을 추출하였다. 그리고, 이들 SNP의 인접서열 왼쪽 500bp와 오른쪽 500bp를 추출하여 SNP를 포함한 1,001 bp의 서열에서 PCR 프라이머를 디자인하였다. 프라이머 디자인은 BatchPrimer3 1.0(http://probes.pw.usda.gov/batchprimer3/) 프로그램을 이용해서 수행하였다. 마커와 마커 사이에 약 2 Mbp 간격을 두고 디자인하였으며, 디자인된 마커로 가야벼와 태백벼의 DNA로 모본간 다형성 검정을 수행하여 실제로 다형성을 보이는 마커들을 선발하였다. 선발된 CAPS 마커들로 F2 집단의 개체별 유전자형 분석을 실시하였다. CAPS 마커 분석의 PCR 조건은 94°C 3분 후 94°C 40초, 60°C 40초, 72°C 1분40초의 35 사이클, 그리고 72°C 7분이었다. PCR 산물을 제한효소로 37°C에서 8시간 이상 절단한 후 1.2% 아가로스젤에서 전기영동하였다. 이렇게 생산된 F2 집단의 개체별 유전자형 데이터를 가지고 MapDisto 1.7 프로그램(Lorieux 2012)을 이용하여 마커들의 유전지도를 작성하였다.
기존에 보고된 벼멸구저항성 유전자 염기서열과 가야벼의 유전체 염기서열 비교
기존에 밝혀진 벼멸구저항성 유전자인 Bph14, Bph3, BPH26, BPH18, BPH9들의 염기서열을 reference 염기서열로 하고, 선행연구(Ji et al. 2016)에서 Illumina Hiseq2000 장비를 이용하여 얻어진 가야벼의 유전체 염기서열들을 reference 염기서열에 read mapping하였다. read mapping은 CLC Assembly Cell 프로그램(ver. 3.2.2, http://www.clcbio.com)의 clc_mapper 명령어를 사용하여 수행하였는데, 이 때 옵션 값을 similarity 98%, matched length fraction 1.0, repeat ignore로 설정하여 각 read의 전체 영역이 match되면서 염기서열 유사도가 98% 이상인 서열만이 reference 염기서열에 mapping 되도록 하고 반복서열들은 제외되도록 하였다. read mapping 결과를 clc_mapping_viewer 명령어를 시용하여 검토함으로써 가야벼의 유전체 염기서열이 비교대상인 각각의 벼멸구저항성 유전자 염기서열과 일치하는지를 판정하였다. 이 결과, 가야벼는 Bph3 유전자와 BPH26 유전자를 가지고 있는 것으로 나타났다. 이를 더 검증하기 위해 가야벼의 Bph3 유전자와 BPH26 영역을 PCR로 증폭하여 Sanger sequencing 방법으로 sequencing 하였다. 이 때 사용한 PCR 프라이머 목록은 Table 1과 같다. Bph3 유전자는 세 개의 lectin receptor kinase 유전자들인 OsLecRK1, OsLecRK2, OsLecRK3로 이루어져 있는데, 이들 각각의 유전자 코딩 시퀀스 영역이 포함되도록 PCR 증폭하여 PCR 산물을 Sanger sequencing 방법으로 sequencing 하였다. 시퀀싱 프라이머는 약 500 bp 간격으로 디자인하여 제작하였다. BPH26 유전자는 크기가 10.6 kbp로 세 개의 엑손(exon)과 두 개의 인트론(intron)으로 이루어져 있는데 Tamura et al. (2014)의 방법으로 엑손 영역만을 PCR 증폭하여 PCR 산물을 Sanger sequencing 방법으로 sequencing 하였다.
Table 1
The list of primers for amplification of Bph3 and BPH26 genes of Gayabyeo
Table 1
   Gene         forward primer         reverse primer gene size (bp)
OsLecRK1 CCCACTCAATTATAGCTACACA TCGACGACAGCAAATCCT 2442
OsLecRK2 ACAAATGCCGTTCTCCAAG AGGATCATGTGGTTGTAGAGG 2436
OsLecRK3 ACGCGGTAGGAAAGACCA AAGCATCAGTGCCCGAACA 2436
BPH26 exon1 TAGCATCAGTCCCTTGCTTGTTTGC ATTGATTTAATTAGCAGACAAGTTG 1089
BPH26 exon2 AACTCTCGTCTCGTCTTAAAATATA AGTAGTAATGTGCGTAGCAATGGAG 624
BPH26 exon3 CTAGTGCCAGTTACTCCGATAAATAT TATGCACTAGCATCACTACA 1944

* Bph3 is composed of three genes that are OsLecRK1, OsLecRK2, and OsLecRK3.

** Sequencing of BPH26 is done on its three exons according to Tamura et al. (2014).

가야벼 저항성 유전자 선발마커 개발
가야벼의 Bph3 유전자와 BPH26 유전자 선발마커를 다음과 같은 방법으로 개발하였다. 가야벼의 유전체 염기서열 데이터를 가지고 CLC Assembly Cell 프로그램의 clc_assembler 명령어를 사용하여 de novo assembly를 수행하였다. 그 결과 생산된 contig 염기서열들을 CLC Genomics Workbench 프로그램(ver. 6.01, http://www.clcbio.com)을 이용하여 BLAST 데이터베이스로 만들고, Bph3와 BPH26 유전자 염기서열로 BLAST를 수행하여 이들 유전자를 포함하는 가야벼 contig들을 추출하였다. 이렇게 추출한 가야벼 contig 염기서열과 자포니카 벼 표준유전체 염기서열인 니폰바레 염기서열(Nipponbare IRGSP-1.0 sequence, http://rapdb.dna.affrc.go.jp/download/irgsp1.html)을 비교하여 삽입/결실(InDel)부위를 찾아내었다. 이들 InDel 부위 양측 인접염기서열을 이용하여 프라이머를 디자인해서 InDel 마커를 개발하였다.
가야벼 벼멸구 저항성 유전분석 집단의 벼멸구 검정 및 QTL 분석
태백벼/가야벼 F2 집단에서 채종한 F3 종자를 이용해서 F3 집단 180 계통에 대한 벼멸구저항성유묘검정을 수행하였다. F3 한 계통당 15립을 파종하여 유묘가 3엽기에 이르렀을 때 2~3령의 벼멸구 유충을 식물체 개체당 10~15마리 되도록 접종하였다. 감수성 대비 품종인 태백벼가 거의 완전히 고사하였을 때 식물체 개체의 저항성지수를 Huang 등에 의해 보고된 기준에 따라 0~9 등급으로 판정하였다(Huang et al. 2001). 개발된 CAPS 마커들 및 Bph3와 BPH26 유전자 선발마커들의 유전지도와 F3 계통별 저항성지수의 평균값을 QTL 분석에 사용하였다. QTL 분석은 Windows QTL Cartographer version 2.5 program(Basten et al. 1996)을 이용하여 composite interval mapping 방법으로 수행하였다. LOD threshold는 확률 수준 0.05에서 1,000번의 permutation을 함으로써 산정되었다.
가야벼와 태백벼간 다형성 SNP 마커 개발
선행연구에서 탐지된 가야벼와 태백벼간 SNP들 가운데 이 연구에서 사용한 제한효소들의 인식부위에 있는 SNP들을 이용하여 CAPS 마커들을 디자인하고 가야벼와 태백벼간 다형성 검정을 하였다. 약 2 Mbp 간격으로 마커들을 디자인 하였으며, 가야벼와 태백벼간 SNP가 드물어서 마커 디자인이 불가능한 영역들도 있었다. 디자인된 마커들로 가야벼와 태백벼 DNA를 이용하여 모본 다형성 검정을 수행하였으며, 모본 품종간 다형성이 확인된 마커들로 태백벼/가야벼 후대 F2 집단 180 개체의 유전자형을 분석하였다. 99 개의 CAPS 마커들이 개발되어, 이들을 이용한 태백벼/가야벼 후대 F2 집단 유전자형 데이터들이 생산되었으며, 이 데이터에 기반하여 개발된 마커들의 유전지도를 작성하였다(Fig. 1). 작성된 유전지도의 총 거리는 1165.5 cM이었고, 마커간 평균 거리는 13.4 cM이었다. 개발된 마커들의 프라이머 염기서열, 제한효소, 물리적 위치를 Table 2에 나타내었다.
Fig. 1
Genetic map of developed CAPS markers.
KJBS_50_104_fig_1.jpg
Table 2
List of developed CAPS markers
Table 2
no. marker name       forward primer       reverse primer restriction enzyme chromosome position (bp)
1 GT01002 GCTGATGATCGTTTGCCACA TGCGATGTATTTTCGCGTTTT EcoRV 1 1,065,395
2 GT01003 AAATTGGCTGGGGCACTTCT TGCAGGATCAGGAAGAGCTG EcoRI 1 1,600,379
3 GT01007 CAGGGCCGCAGTGTTAATTC AGCGAGACGTTCGTGATGGT EcoRI 1 5,522,341
4 GT01008 AATAAGCGCTTGACGCGAAA CGCTCAACCACCTCGTCATT EcoRV 1 6,037,347
5 GT01011 TCCTGAGCATCCTTGCATGA AAAACGAGAGGGCGCATGT EcoRV 1 9,187,583
6 GT01018 ACTTGGAGGCGCATAACGAA GCATAATGACATGTCGCATTGAA EcoRI 1 23,956,461
7 GTS01007 AAGGGTTGTATGAACCGTCAAC TCCATGGGACACTAAAGAGAGG SalI 1 25,601,011
8 GTS01008 CGTCGAAGAAAGTGACTCAAGA CCTCTTTTGTTGGTATCATTGG NruI 1 26,694,741
9 GTS01009 ACTCCCAAAACCAGCGTAAATA CGTCAGGAATGAATTCGAAACT SacI 1 27,335,280
10 GTS01010 AGAACCCGTGTAGAGCACATTA CAATTGATCTGCTTGCTCTCTG PshBI 1 32,523,034
11 GT02001 GCATATGTTTGACCGTTCGTCTT CGGCGCTACACCATACCAAT EcoRI 2 198,216
12 GTS02001 AACTAGCACGCTTCGTTTTAGC CGGATTGGTGGAATTGAAAA StuI 2 1,330,779
13 GTS02002 CCATTTGGCAATGTTCTCATAC CACCCTTCTATGCTCTCATCCT BamHI 2 4,001,036
14 GTS02003 TGTGTGGTAGAAAATGGACCTG CCATGATTGCCAGTGTTACATC EcoRI 2 5,066,630
15 GTS02004 TACCGATCAGCTGAAAACACAC TCTTGTTCCCTTCCCTTTAACA BamHI 2 6,854,129
16 GTS02006 GTGACCTCCGATCAAATCAACT CTACTGTGGGCAGTGAAGAGGT BamHI 2 8,587,683
17 GTS02007 ATTTCCCTCCTGTTCCTCATTT GGTTGGGTTTTGGTATAGCTGA HindIII 2 9,657,613
18 GTS02008 AGCTGGAAAATGTTGTATGCAG GGGATGAATCAGTAGTTCAGCA HindIII 2 10,645,645
19 GT02014 GTGTTGGCTAGGGCCTTTGA TTCTCTGGTTGGCCTTCAGC EcoRI 2 21,056,727
20 GT02021 CGCCTCTTGAAGACCCAGAG CTGACCGGCATGGGAGTTTA SacI 2 33,887,583
21 GT03001 GCTGTTGCCGCAATATCACA AGGCATTGGCATTGTTAGGC EcoRI 3 855,210
22 GT03003 TGGGGATTGCCAGATGTGAT GCATGGCATTTGAAAGAGTGA PvuII 3 2,538,138
23 GT03004 CAGTGTCTGGGCACAGCAAG GCGTTCCACACTTCCACTCC HindIII 3 3,811,855
24 GT03005 GGAAATACCTGCCCCGTTTT CAGACGATCGGGAATTTCTACG EcoRV 3 4,753,112
25 GT03006 CTCCTCTCGCACGCAGAAGT CCAGCTTCTCCACGGAACAA EcoRV 3 5,592,985
26 GT03009 TCTACCTGCCCAGGAAGCAA TTTCAGGCAAAGCAAAGCAG EcoRV 3 9,205,476
27 GTS03002 AGCATCACGATGATTCTTTTGC TTATTGTGCCCTAGACGAATGA SacI 3 11,547,474
28 GTS03004 TAGGGTTGGCTCTCAATCTCTC ACACCTACAAGGTTGTGACTGC EcoRI 3 13,593,894
29 GTS03008 ATCGCACGGAAGGATAACTG GTTGCTCTAGAACCTCGCAATC SacII 3 19,416,808
30 GTS03009 ATTATCTCACGACCTGGGACTG AGCTAGCTAGGGTTTGTGTTGC EcoRI 3 20,345,341
31 GTS03010 TGAGGAAGAGGAGGAGATTGAG CTGCTTCATTGTGTCTGAGGAC SacI 3 21,733,967
32 GT03021 GAATCTGATTGCCCCCTAGC GGACCCAAATTGAAAGGCAAT EcoRV 3 25,459,877
33 GT03024 TTGGAAGCTTGAAACGATGG TCGCAAGTTCGAAACCCACT EcoRI 3 28,056,489
34 GT03025 TGCGAATACAATTCGGACTGC CGTGCAGTACCCTCCAAACC EcoRV 3 29,184,626
35 GT03030 AATTAAACTCCGCGCCTGGT AAACCATGCATCCGATAGGG HindIII 3 36,245,706
36 GT04004 TGGCGACAAACAGAGCACAC CCTTCTTGCGCAGGACCTC EcoRI 4 4,875,618
37 GT04005 CCAGCCAAACTCAGTTGTCCA CAGACCATACCTCCTTCAGAGCA EcoRI 4 5,924,429
38 GTS041424 ACTGGGAAGAAGGCAAGACA GTGAGCATCCCACTCTCCAT EcoRI 4 6,623,655
39 GTS041472 CCTGAAAGGGCTCAAAAACA TCTTGCAGGTGACAAATCCA PshBI 4 6,914,922
40 GTS041749 TGAACTATTGCGGTTGACCA TGCGGATTGTGTAAGACAGG EcoRV 4 8,481,173
41 GT04014 TGGATCGCTTGATGTGGTTG ATGCCTCAAATACCGCAGCA EcoRI 4 14,685,347
42 GTS043975 TTCGAGGGTTGGTGACTGAT AGAGGGTCCATGGTTTTGTG EcoRV 4 16,607,468
43 GT04020 TCCTTGCAAACGCAAAGTCA GCAACAAAACATTGCGAAGG EcoRI 4 21,083,274
44 GT05001 TTGCAGCTGAAGAAAATGTG TTTGGCACATGAAAATGATG EcoRI 5 602,152
45 GT05003 TGTTGCAGAGTAGCAAACCA TATAAGCTGCAGCCTGGAAT EcoRV 5 2,107,453
46 GTC05003 CCGAAGAGCGACTCCTAGTG AGAGATGCACAGAGGGAGGA PshBI 5 6,043,943
47 GT05013 GAGCATAAGGGGTTTCCCTA TAATGCTGGTGTGGACTCAA EcoRI 5 15,946,504
48 GTC05008 TAATTGCCTCCCAAACAACC TGTCATGCAAAGCAAAAAGG PshBI 5 18,530,587
49 GT05015 CGACGACTATATGGCGGATA AACTCCAATCGGTCATACCA EcoRI 5 20,548,891
50 GT05016 CCGTTTTGTCAGAAACCTGT TAAATCGCGAGACGAATCTT EcoRV 5 22,331,098
51 GTC05010 AGGTGGTGGTGGTTCCATT TCACTGCCGATTTCTCACTG PvuII 5 25,145,098
52 GT05024 TGATCCCCAATGAAAGCTAC TCTTGTACTCCCTTCGTTTCA EcoRI 5 29,110,069
53 GT06013 TCCAACAGCAACAATCTTCA TCTCAAGCTCAATTCCGAGT EcoRI 6 24,403,695
54 GT06015 AGAGCAGAGCTCGTGTTAGTG TGGTGATTTTTGGATGGATT EcoRI 6 26,356,825
55 GT06016 GCGAAGATGCTAGTGGAAAA GTTGCAAAGGATCTGGAAAA EcoRV 6 27,324,979
56 GT06017 GATTCGACGTCTGAAAAGGA ATATGCATGGTTCACTGCAA EcoRI 6 29,168,500
57 GT06019 TGCTCAGCAGAGTCTTGCTAT GCTTGAGACTGGTTCGTGTT SacII 6 31,015,468
58 GT07002 CAAAAGTAACCAATGGCAAGA GCACATTTGAACTGCATCTG EcoRV 7 5,619,862
59 GT07008 TGGAATCAAAGTGAAGGTCAG TGGCAATCTAAAGAGGAGGA EcoRI 7 17,797,502
60 GT07010 TATGGGGACTCCAATAAGCA TACGGTCTCGTCGGTATCAT EcoRV 7 19,489,327
61 GTS07491 AAGGCCAAATGGAGGTTAGC CTGGGAGTCCAGTCATCACA BglII 7 21,220,123
62 GT07013 ATTCAGCCCCTTCTGGATAC ACCTAGGGCAGTCCTTGATT EcoRI 7 22,517,510
63 GTS07984 GCCGCTTTCTAAAAGGATTT GACCGAAATTTTGGCTCAAT EcoRV 7 23,387,924
64 GTS07117 AGCCTGCCACTCTTCCTGTA CGAGGACATGGCATCCTATT EcoRV 7 27,812,959
65 GT07018 TTCAACCTTAGGCACTCGTC TGAGATTCTCCTTGCGTGTT EcoRV 7 28,923,650
66 GTS08007 GATCAGTCGTCCTCGTCCTC GCATTGAGCACATCCATGAC HindIII 8 1,150,395
67 GTS08142 ATGCTCCAATTGAGCTGACC TACTCATGCAGGAACGCATC EcoRI 8 6,123,634
68 GTS08218 AAATGGTGTTTTTCCCGTTG AGGTCGTGGCATGATTTAGC PshBI 8 8,787,812
69 GTS08298 CGATCGGCTCTTACATGACA AGTGGATTTGCAGGCAGCTA EcoRI 8 27,252,203
70 GTS08328 GAGTTACAAACCGCCTTTGC TTGCTAGGGGTCAAGGAAAA BglII 8 28,243,051
71 GTS09004 AGCACCCAGAGCAACAGTTT CCCACTAATTAGCCCATTGC EcoRV 9 620,373
72 GTS09063 GCTTGGATGACAAGCGTTCT AGATCGGCTTCCGAAAGAGT BglII 9 9,835,918
73 GTS09106 ACGCGACTGCTAAGGTTCAT CTCGATCGGTCCCTTTCAT EcoRV 9 11,310,635
74 GTS09188 AGTGGTAGATCCCGTTGAGC AACCGGAGACCTTCAGTGTG EcoRV 9 13,099,244
75 GTS09302 GCATGCATCCGTTGAATAAA GAGATGCTCGTCAGGGACAC HindIII 9 16,712,838
76 GTS09305 ATGGCTTTCGGGCATAGTAA GGATGGCCTATTCAGCAAAA EcoRV 9 21,161,450
77 GTS10005 TCTCAGCCACTCGTTGAATG TCTGGATGGGCTAAATGAGG EcoRV 10 721,349
78 GTS10029 CCTCCCTTCAGTTTGTGAGC CATTGGATCCCCAAAGAAAA EcoRI 10 3,044,726
79 GTS10116 CCCCCTAAGATGGTTTTTGC TGAGAAGTGAAGTGTGAGTGTGG EcoRV 10 4,577,975
80 GTS10199 AGCCTGAGAAGCAGATTGGA TGCAATGACGTGATGGTACA EcoRI 10 6,149,421
81 GTS10204 TTTCACACGGGACATTCAAA TCGACCTTGGACTTCTAGGC StuI 10 9,961,760
82 GTS10214 TAGGATCTCGCCGAACATCT ATGCACCTCCCCAACATATC EcoRV 10 11,290,285
83 GTS10276 TGGTCTTTGAAGCATTCTGG TTCAATATTGGCCGTAAACC EcoRV 10 13,637,268
84 GTS11014 CTTCGCTTCATTTCGAGTTTG CATTTCTGCAGTCGCATTTG EcoRV 11 353,397
85 GTS11016 CGCACGAGATGCTGTCTAAC GAAAAACCCTCTGCCCTCTT NruI 11 1,389,204
86 GTS11086 TGCTGGTGTGCTATTTGAGC CGTGGCCATCCAAAAGTAAT EcoRI 11 17,330,622
87 GTS11088 CGCCTAGGTGCAGAAAAGAG ACACCATGAGCCTCCTCACT BamHI 11 20,838,960
88 GTS11330 TGTCATTGGTTGCCCTGTAA AGCATGGAGGTGGTGCTAAC EcoRV 11 26,534,991
89 GTS11350 AGGAATATGAATCGCGTGTG TTTGGGAGCTATGGCCTATG EcoRI 11 27,197,820
90 GTS120003 GCGTTTTTCCTCCAATTCAA ACCGACAACTTTTGCCCTTT PvuII 12 3,007,338
91 GTS120032 AACGAAGAGGCCAATGGATA GTTGACCATGGGTGCTCACT PvuII 12 3,807,192
92 GTS120139 TGCTTTCAGGAGAGCAGGAT CCACCAGCATCACTCTGCTA EcoRI 12 9,263,934
93 GTS120457 GCATGAGAGACAGCGGAGTT GTGATTTGCCATGCCTTTTT EcoRV 12 13,674,598
94 GTS120652 TGGGCACAACTACAAAGGTG CATTCCCATGTTCCACATCA EcoRI 12 17,438,893
95 GTS120760 CCCAGCCACAAGAAATGAAT TTTTCCTTTTCCCTGCTGTG EcoRI 12 19,070,762
96 GTS120842 AAGCCATCATGTTCCATTCC TTTTGGTACGTCATCCGTGT EcoRI 12 21,522,146
97 GTS120941 CAGGTGGTCCTTTTCAGCAT TAGTGTCATGTGCCCTGAGC EcoRI 12 22,565,939
98 GTS120989 CCTTTGGAGGGCTTGACATA TACCCACTGGAAACGGAAGA EcoRV 12 23,622,982
99 GTS121030 CGTTGCTCCGTTCATCTTCT GCTAGCCCCTTTTCATCTCC EcoRV 12 26,276,873
유전체 염기서열 데이터 분석에 의한 가야벼 벼멸구 저항성 유전자 규명
기존에 밝혀진 벼멸구저항성 유전자인 Bph14, Bph3, BPH26, BPH18, BPH9들의 염기서열을 reference 염기서열로 하여, 가야벼의 유전체 염기서열을 read mapping하여 비교한 결과, 가야벼의 유전체 염기서열이 Bph3과 BPH26 유전자 염기서열과 일치하였다(Fig. 2). Fig. 2에서 reference 염기서열과 일치하는 유전체 염기서열은 흰색으로 나타나고, 일치하지 않는 염기서열은 적색으로 나타나는데, Bph3과 BPH26 유전자에서는 전 영역에 걸쳐 염기서열이 일치한 반면 Bph14, BPH18, BPH9 유전자에서는 일치하지 않는 염기서열이 다수 존재하였고, read mapping이 되지 않는 영역이 존재하였다. 따라서, 가야벼가 Bph3과 BPH26 저항성 유전자를 가지고 있다고 판단되었다.
Fig. 2
Read mapping of Gayabyeo genome sequences onto BPH resistance genes.
Reference BPH resistance gene sequences were shown at the top row of each panel, and mapped read sequences were shown below the reference sequences. White letters indicate identical nucleotides while red letters indicate different nucleotides. a: Bph3, b: BPH14, c: BPH18, d: BPH26, e: BPH9.
KJBS_50_104_fig_2.jpg
이 결과를 검증하기 위해 가야벼의 Bph3과 BPH26 유전자 영역을 Table 1의 프라이머로 PCR 증폭하여 Sanger sequencing 방법으로 sequencing하여 Bph3과 BPH26 유전자 염기서열과 비교하였다. Bph3 유전자를 구성하는 세 개의 lectin receptor kinase 유전자들인 OsLecRK1, OsLecRK2, OsLecRK3 각각의 가야벼 PCR 산물의 염기서열이 Bph3 유전자 염기서열과 완전히 일치하였다. 그리고, 가야벼의 BPH26 유전자 세 개 엑손 영역 PCR 산물의 염기서열도 BPH26 유전자 영역 염기서열과 완전히 일치하였다. 이로써, 가야벼가 Bph3과 BPH26 저항성 유전자를 가지고 있음을 확인할 수 있었다.
가야벼 저항성 유전자 선발 마커 개발
가야벼가 벼멸구 저항성 유전자 Bph3과 BPH26을 가지고 있으므로, 이들 유전자들을 국내 자포니카형 벼 품종들에 도입하는 육종프로그램에 사용할 수 있는 선발마커를 개발하였다. 특히, 일반적인 실험실에서 아가로스 젤을 이용하여 쉽게 분석할 수 있는 삽입/결실(InDel) 마커를 개발하고자 하였다. 가야벼의 유전체 염기서열과 자포니카형 벼 품종들의 유전체 염기서열간 InDel 부위를 찾기 위해서, 먼저 가야벼의 유전체 염기서열 데이터로 de novo assembly를 수행하여 contig 들을 만들었다. de novo assembly 결과 200~95,218 bp 범위의 크기를 가진 contig들이 64,373개 생성되었으며, BLAST를 통해서 Bph3유전자와 BPH26 유전자 영역에 있는 가야벼 contig들을 추출하였다. 추출된 가야벼 contig들의 sequence와 자포니카 벼 표준유전체 염기서열인 니폰바레 염기서열과의 비교를 통해서 InDel 부위를 발굴하였다. Bph3유전자를 구성하고 있는 OsLecRK1, OsLecRK2, OsLecRK3 유전자 영역에서 OsLecRK1 유전자의 3’ 말단 인접부위에서 가야벼 contig 14558과 니폰바레 유전체 염기서열간 72 bp InDel 부위를 발굴하고(Fig. 3a) InDel 마커 Bph3IND를 개발하였다(Table 3). BPH26유전자는 세개의 exon과 두 개의 intron으로 구성되어 있는데 두번째 intron에서 가야벼 contig 36193과 니폰바레 유전체 염기서열간 587 bp InDel 부위를 발굴하고(Fig. 3b) InDel 마커 Bph26IND를 개발하였다(Table 3). 이들 마커들로 가야벼와 34개의 국내육성 자포니카 벼 품종들의 유전자형을 분석한 결과, Bph3IND 마커와 Bph26IND 마커 둘 다 가야벼와 국내 자포니카 품종들간에 뚜렷한 차이를 보였다(Fig. 4a). 그러므로 이 마커들은 가야벼의 벼멸구 저항성 유전자들을 국내 자포니카 벼 품종들에 도입하는 여교배 육종 프로그램에 선발마커로 유용하게 사용될 수 있을 것으로 판단된다. 한편, Bph3IND 마커로 가야벼와 15개의 국내육성 통일형 벼 품종들의 유전자형을 분석한 결과, Bph3IND 마커에서는 남영벼만이 가야벼와 동일한 밴드를 보였고, 태백벼 등 5품종에서 가야벼와 다른 밴드가 관찰되었으며, 밀양23호 등 9 품종에서는 밴드가 관찰되지 않았다(Fig. 4b). 이로써 벼멸구 저항성 품종으로 알려진 남영벼가 Bph3 유전자를 가지고 있을 가능성이 있다고 보여진다. 또한, Bph26IND 마커로 가야벼와 15개의 국내육성 통일형 벼 품종들의 유전자형을 분석한 결과, 밀양23호 등 12 품종에서는 가야벼와 다른 밴드가 관찰되었고, 삼강벼, 안다벼, 용문벼에서는 가야벼와 동일한 밴드가 관찰되었으나 밴드가 두 개인 hetero type들이어서 가야벼와 다른 양상을 보였다. 이로써, 국내육성 통일형 벼 품종들에서도 Bph3IND 마커와 Bph26IND 마커의 활용가능성이 있다고 판단된다.
Fig. 3
Detection of InDels between Gayabyeo and Nipponbare reference sequences for development of InDel markes in Bph3 and BPH26 region. a: Bph3 region, b: BPH26 region.
KJBS_50_104_fig_3.jpg
Table 3
Primer sequences of Bph3IND and BPH26IND markers.
Table 3
marker name      forward primer      reverse primer
Bph3IND GCTTCACCTGTGTCCTGTT GTGTATGAGGGTTGTTCTGGAT
BPH26IND GGGCAGCTAAGCGCATGGTT GGGTGATAAATCGAACAGCAGAGG
Fig. 4
Genotyping of Gayabyeo and other Korean rice varieties with Bph3IND and BPH26IND markers.
a: genotyping Gayabyeo and 34 Korean japonica rice varieties, b: genotyping Gayabyeo and 15 Korean Tongil-type rice varieties.
KJBS_50_104_fig_4.jpg
가야벼의 벼멸구저항성 QTL mapping
태백벼/가야벼 F3 집단 180 계통에 대해서 벼멸구저항성 유묘검정을 수행한 결과, F3 계통들의 계통별 저항성 지수 평균값은 Fig. 5과 같은 분포를 보였다. F3 계통들의 계통별 저항성 지수 평균값과 개발된 CAPS 마커 및 Bph3IND 마커와 Bph26IND 마커로 작성한 유전지도를 종합하여 저항성 QTL을 분석하였다. 그 결과 4번 염색체 0 - 6.6 cM 영역에서 LOD 값이 7.22인 QTL인 qBPH4가 발견되었고, 12번 염색체 40.8 – 54.4 cM 영역에서 LOD 값이 6.64인 QTL인 qBPH12가 발견되었다(Table 4). 확률 수준 0.05에서 1,000번의 permutation을 함으로서 결정된 LOD threshold 값은 3.9이었다. qBPH4 영역은 Bph3IND 마커(5.7 cM)을 포함하고 있었고, qBPH12 영역은 BPH26IND 마커(48.4 cM)을 각각 포함하고 있었다. 따라서, qBPH4는 Bph3 유전자이며, qBPH12는 BPH26 유전자로 판단되며, 이 결과는 가야벼의 벼멸구 저항성 유전자가 Bph3와 BPH26임을 뒷받침해주고 있다고 판단된다.
Fig. 5
Distribution of BPH resistance scores of the 180 F3 families.
KJBS_50_104_fig_5.jpg
Table 4
Identification of QTLs for BPH resistance
Table 4
QTL name chromosome location (cM) QTL interval* (cM) LOD additive effect dominance effect R2
 qBPH4 4 0 0-6.6 7.22 0.94 -0.20 0.138
 qBPH12 12 51.4 40.8-54.4 6.64 0.87 0.55 0.131

*interval at 99% probability

통일형 벼품종인 가야벼는 우리나라에서 수집된 다양한 벼멸구 집단들에 대해서 저항성을 가진 것으로 알려져 있다. 가야벼 벼멸구 저항성의 유전분석을 위해서, 가야벼와 벼멸구 감수성 통일형 품종인 태백벼를 교배하여 F2 및 F3 집단을 작성하였다. 그리고, 선행연구에서 탐색된 가야벼와 태백벼간 약 28만개의 단일염기서열변이(SNPs)들 중 제한효소 인식부위에 있는 SNP들을 대상으로 99 개의 CAPS (Cleaved Amplified Polymorphic Sequences) 마커를 개발하였다. 이들 마커들로 태백벼/가야벼 F2 집단 180 개체들의 유전자형을 분석하여 유전지도를 작성하였다. 가야벼의 유전체재해석(resequencing) 염기서열 데이터와 기존에 알려진 벼멸구 저항성 유전자들(Bph3, BPH9, Bph14, BPH18, BPH26)의 염기서열을 비교한 결과, 가야벼가 Bph3과 BPH26 유전자들을 가지고 있는 것으로 나타났다. 가야벼의 Bph3과 BPH26 유전자 영역을 PCR 증폭하여 Sanger sequencing 방법으로 염기서열을 분석한 결과, Bph3 및 BPH26 유전자 염기서열과 완전히 일치함을 확인하였다. 이 두 유전자 영역에서 삽입/결실(InDel) 마커인 Bph3IND와 BPH26IND를 개발하였고, 이들 마커들은 가야벼의 저항성 유전자들을 고품질 자포니카 벼 품종들에 도입하는 육종 프로그램에 선발마커로서 활용될 수 있음을 확인하였다. 태백벼/가야벼 F3 집단 180 계통들을 대상으로 벼멸구저항성 유묘검정을 수행하여 QTL 분석을 한 결과, 4번 염색체와 12번 염색체에서 주동 QTL 두 개가 발견되었다. 이 두 QTL들의 영역은 각각 Bph3과 BPH26 유전자를 포함하고 있었는데, 이는 가야벼가 Bph3과 BPH26 유전자를 가지고 있음을 뒷받침해 주었다. 이 결과는 우리나라에서 고품질 벼멸구저항성 자포니카 벼 품종 육성을 촉진하는데 유용하게 활용될 수 있을 것이다.
본 논문은 농촌진흥청 연구사업(과제번호: PJ01104201)의 지원에 의해 이루어진 것임.
  • Basten CJ, Weir BS, Zeng ZB. 1996. QTL Cartographer: A reference manual and tutorial for QTL mapping. Department of Statistics, North Carolina State University. Raleigh, NC.
  • Cantindig JLA, Arida GS, Baehaki SE, Bentur JS, Cuong LQ, Norowi M, Rattanakam W, Sriratanasak W, Xia J, Lu Z. 2009. Situation of planthoppers in Asia. Heong KL, Hardy B, editors. Planthoppers: new threats to the sustainability of intensive rice production systems in Asia. International Rice Research Institute. Los Baños, Philippines: pp. 191-220.
  • Du B, Zhang W, Liu B, Hu J, Wei Z, Shi Z, He R, Zhu L, Chen R, Han B, He G. 2009. Identification and characterization of Bph14 a gene conferring resistance to brown planthopper in rice. Proc Natl Acad Sci U S A 106: 22163-22168.
  • Fujita D, Kohli A, Horgan FG. 2013. Rice Resistance to Planthoppers and Leafhoppers. Critical Reviews in Plant Sciences 32: 162-191.
  • Hibino H. 1996. Biology and epidemiology of rice viruses. Annu Review of Entomology 34: 249-274.
  • Hu J, Cheng MX, Gao GJ, Zhang QL, Xiao JH, He Y. 2013. Pyramiding and evaluation of three dominant brown planthopper resistance genes in the elite indica rice 9311 and its hybrids. Pest Management Science 69: 802-808.
  • Hu J, Xiao C, He Y. 2016. Recent progress on the genetics and molecular breeding of brown planthopper resistance in rice. Rice (N Y) 9: 30
  • Huang Z, He G, Shu L, Li X, Zhang Q. 2001. Identification and mapping of two brown planthopper resistance genes in rice. Theor Appl Genet 102: 929-934.
  • Ji H, Ahn E, Seo BY, Kang HJ, Choi I, Kim KH. 2016. Genome-wide detection of SNPs between two Korean Tongil-type rice varieties. Korean J Breed Sci 48: 460-469.
  • Ji H, Kim SR, Kim YH, Suh JP, Park HM, Sreenivasulu N, Misra G, Kim SM, Hechanova SL, Kim H, Lee GS, Yoon UH, Kim TH, Lim H, Suh SC, Yang J, An G, Jena KK. 2016. Map-based cloning and characterization of the BPH18 gene from wild rice conferring resistance to brown planthopper (BPH) insect pest. Sci Rep 6: 34376
  • Kim DH, Lee GH, So JD, Yuh JH, Kim JH. 1991. Resistance aspects on different growth stages and field resistance of rice varieties to brown planthopper Nilaparvata lugens STAL (Homoptera : Delphacidae). Research Report of RDA 33: 33-39.
  • Kim YH, Lee JO, Choi KM, Cho SY. 1989. Varietal resistance of Korea rice cultivars to brown planthopper and whitebacked planthopper and their resistance sources. Research Report of RDA 31: 7-13.
  • Lee JO, Goh HG, Kim YH. 1985. Study on brown planthopper biotypes. 2. Resistance mechanism of different rice varieties to Korean brown planthopper biotypes. Research Report of RDA 27: 73-78.
  • Lee JO, Goh HG, Kim HS, Choi KM. 1986. Study on brown planthopper biotypes. 3. Plant damage of different rice cultivars to combination of biotypes. Research Report of RDA 28: 52-55.
  • Liu Y, Wu H, Chen H, Liu Y, He J, Kang H, Sun Z, Pan G, Wang Q, Hu J, Zhou F, Zhou K, Zheng X, Ren Y, Chen L, Wang Y, Zhao Z, Lin Q, Wu F, Zhang X, Guo X, Cheng X, Jiang L, Wu C, Wang H, Wan J. 2014. A gene cluster encoding lectin receptor kinases confers broad-spectrum and durable insect resistance in rice. Nat Biotechnol 33: 301-305.
  • Lorieux M. 2012. MapDisto: fast and efficient computation of genetic linkage maps. Mol Breeding 30: 1231-1235.
  • Myint KK, Fujita D, Matsumura M, Sonoda T, Yoshimura A, Yasui H. 2012. Mapping and pyramiding of two major genes for resistance to the brown planthopper (Nilaparvata lugens [Stal]) in the rice cultivar ADR52. Theor Appl Genet 124: 495-504.
  • Otuka A. 2009. Migration of rice planthoppers and simulation techniques. Heong KL, Hardy B, editors. Planthoppers: new threats to the sustainability of intensive rice production systems in Asia. International Rice Research Institute. Los Baños, Philippines: pp. 221-230.
  • Park YD, Song YH. 1988. Preference, development and fecundity of the brown planthopper (Nilaparvata lugens Stal) biotypes fed on different cultivars of rice with various resistance gene. Korean Journal of Applied Entomology 27: 87-93.
  • Qiu Y, Guo J, Jing S, Zhu L, He G. 2012. Development and characterization of japonica rice lines carrying the brown planthopper-resistance genes BPH12 and BPH6. Theor Appl Genet 124: 485-494.
  • Seo BY, Jung JK, Choi BR, Park HM, Lee BH. 2009. Resistance-breaking ability and feeding behavior of the brown planthopper Nilaparvata lugens recently collected in Korea. Heong KL, Hardy B, editors. Planthoppers: new threats to the sustainability of intensive rice production systems in Asia. International Rice Research Institute. Los Baños, Philippines: pp. 303-314.
  • Seo BY, Kwon YH, Jung JK, Kim GH. 2009. Electrical penetration graphic waveforms in relation to the actual positions of the stylet tips of Nilaparvata lugens in rice tissue. Asia-Pacific Entomology 12: 89-95.
  • Sogawa K. 1982. The rice brown planthopper: feeding physiology and host plant interactions. Annual Review of Entomology 27: 49-73.
  • Tamura Y, Hattori M, Yoshioka H, Yoshioka M, Takahashi A, Wu J, Sentoku N, Yasui H. 2014. Map-based cloning and characterization of a brown planthopper resistance gene BPH26 from Oryza sativa L. ssp. indica cultivar ADR52. Sci Rep 4: 5872
  • Uhm KB, Park JS, Lee YI, Choi KM, Lee MH, Lee JO. 1988. Relationship between some weather conditions and immigration of the brown planthopper Nilaparvata lugens Stål. Koream Journal of Applied Entomology 27: 200-210.
  • Wilson MR, Claridge MF. 1991. Handbook for the identification of leafhoppers and planthoppers of rice. CAB International. Wallingford, UK.
  • Zhao Y, Huang J, Wang Z, Jing S, Wang Y, Ouyang Y, Cai B, Xin XF, Liu X, Zhang C, Pan Y, Ma R, Li Q, Jiang W, Zeng Y, Shangguan X, Wang H, Du B, Zhu L, Xu X, Feng YQ, He SY, Chen R, Zhang Q, He G. 2016. Allelic diversity in an NLR gene BPH9 enables rice to combat planthopper variation. Proc Natl Acad Sci U S A 113: 12850-12855.

Download Citation

Download a citation file in RIS format that can be imported by all major citation management software, including EndNote, ProCite, RefWorks, and Reference Manager.

Format:

Include:

Detection of Genes Conferring Resistance to the Brown Planthopper (BPH) in Gayabyeo Through Genome Resequencing and Development of Their Selection Markers
Korean. J. Breed. Sci.. 2018;50(2):104-115.   Published online June 1, 2018
DOI: https://doi.org/10.9787/KJBS.2018.50.2.104
Download Citation

Download a citation file in RIS format that can be imported by all major citation management software, including EndNote, ProCite, RefWorks, and Reference Manager.
Format:
Include:
Detection of Genes Conferring Resistance to the Brown Planthopper (BPH) in Gayabyeo Through Genome Resequencing and Development of Their Selection Markers
Korean. J. Breed. Sci.. 2018;50(2):104-115.   Published online June 1, 2018
DOI: https://doi.org/10.9787/KJBS.2018.50.2.104
Close

Figure

  • 0
  • 1
  • 2
  • 3
  • 4
Detection of Genes Conferring Resistance to the Brown Planthopper (BPH) in Gayabyeo Through Genome Resequencing and Development of Their Selection Markers
Image Image Image Image Image
Fig. 1 Genetic map of developed CAPS markers.
Fig. 2 Read mapping of Gayabyeo genome sequences onto BPH resistance genes. Reference BPH resistance gene sequences were shown at the top row of each panel, and mapped read sequences were shown below the reference sequences. White letters indicate identical nucleotides while red letters indicate different nucleotides. a: Bph3, b: BPH14, c: BPH18, d: BPH26, e: BPH9.
Fig. 3 Detection of InDels between Gayabyeo and Nipponbare reference sequences for development of InDel markes in Bph3 and BPH26 region. a: Bph3 region, b: BPH26 region.
Fig. 4 Genotyping of Gayabyeo and other Korean rice varieties with Bph3IND and BPH26IND markers. a: genotyping Gayabyeo and 34 Korean japonica rice varieties, b: genotyping Gayabyeo and 15 Korean Tongil-type rice varieties.
Fig. 5 Distribution of BPH resistance scores of the 180 F3 families.

Fig. 1

Fig. 2

Fig. 3

Fig. 4

Fig. 5

Detection of Genes Conferring Resistance to the Brown Planthopper (BPH) in Gayabyeo Through Genome Resequencing and Development of Their Selection Markers

The list of primers for amplification of Bph3 and BPH26 genes of Gayabyeo

   Gene         forward primer         reverse primer gene size (bp)
OsLecRK1 CCCACTCAATTATAGCTACACA TCGACGACAGCAAATCCT 2442
OsLecRK2 ACAAATGCCGTTCTCCAAG AGGATCATGTGGTTGTAGAGG 2436
OsLecRK3 ACGCGGTAGGAAAGACCA AAGCATCAGTGCCCGAACA 2436
BPH26 exon1 TAGCATCAGTCCCTTGCTTGTTTGC ATTGATTTAATTAGCAGACAAGTTG 1089
BPH26 exon2 AACTCTCGTCTCGTCTTAAAATATA AGTAGTAATGTGCGTAGCAATGGAG 624
BPH26 exon3 CTAGTGCCAGTTACTCCGATAAATAT TATGCACTAGCATCACTACA 1944

* Bph3 is composed of three genes that are OsLecRK1, OsLecRK2, and OsLecRK3.

** Sequencing of BPH26 is done on its three exons according to Tamura et al. (2014).

List of developed CAPS markers

no. marker name       forward primer       reverse primer restriction enzyme chromosome position (bp)
1 GT01002 GCTGATGATCGTTTGCCACA TGCGATGTATTTTCGCGTTTT EcoRV 1 1,065,395
2 GT01003 AAATTGGCTGGGGCACTTCT TGCAGGATCAGGAAGAGCTG EcoRI 1 1,600,379
3 GT01007 CAGGGCCGCAGTGTTAATTC AGCGAGACGTTCGTGATGGT EcoRI 1 5,522,341
4 GT01008 AATAAGCGCTTGACGCGAAA CGCTCAACCACCTCGTCATT EcoRV 1 6,037,347
5 GT01011 TCCTGAGCATCCTTGCATGA AAAACGAGAGGGCGCATGT EcoRV 1 9,187,583
6 GT01018 ACTTGGAGGCGCATAACGAA GCATAATGACATGTCGCATTGAA EcoRI 1 23,956,461
7 GTS01007 AAGGGTTGTATGAACCGTCAAC TCCATGGGACACTAAAGAGAGG SalI 1 25,601,011
8 GTS01008 CGTCGAAGAAAGTGACTCAAGA CCTCTTTTGTTGGTATCATTGG NruI 1 26,694,741
9 GTS01009 ACTCCCAAAACCAGCGTAAATA CGTCAGGAATGAATTCGAAACT SacI 1 27,335,280
10 GTS01010 AGAACCCGTGTAGAGCACATTA CAATTGATCTGCTTGCTCTCTG PshBI 1 32,523,034
11 GT02001 GCATATGTTTGACCGTTCGTCTT CGGCGCTACACCATACCAAT EcoRI 2 198,216
12 GTS02001 AACTAGCACGCTTCGTTTTAGC CGGATTGGTGGAATTGAAAA StuI 2 1,330,779
13 GTS02002 CCATTTGGCAATGTTCTCATAC CACCCTTCTATGCTCTCATCCT BamHI 2 4,001,036
14 GTS02003 TGTGTGGTAGAAAATGGACCTG CCATGATTGCCAGTGTTACATC EcoRI 2 5,066,630
15 GTS02004 TACCGATCAGCTGAAAACACAC TCTTGTTCCCTTCCCTTTAACA BamHI 2 6,854,129
16 GTS02006 GTGACCTCCGATCAAATCAACT CTACTGTGGGCAGTGAAGAGGT BamHI 2 8,587,683
17 GTS02007 ATTTCCCTCCTGTTCCTCATTT GGTTGGGTTTTGGTATAGCTGA HindIII 2 9,657,613
18 GTS02008 AGCTGGAAAATGTTGTATGCAG GGGATGAATCAGTAGTTCAGCA HindIII 2 10,645,645
19 GT02014 GTGTTGGCTAGGGCCTTTGA TTCTCTGGTTGGCCTTCAGC EcoRI 2 21,056,727
20 GT02021 CGCCTCTTGAAGACCCAGAG CTGACCGGCATGGGAGTTTA SacI 2 33,887,583
21 GT03001 GCTGTTGCCGCAATATCACA AGGCATTGGCATTGTTAGGC EcoRI 3 855,210
22 GT03003 TGGGGATTGCCAGATGTGAT GCATGGCATTTGAAAGAGTGA PvuII 3 2,538,138
23 GT03004 CAGTGTCTGGGCACAGCAAG GCGTTCCACACTTCCACTCC HindIII 3 3,811,855
24 GT03005 GGAAATACCTGCCCCGTTTT CAGACGATCGGGAATTTCTACG EcoRV 3 4,753,112
25 GT03006 CTCCTCTCGCACGCAGAAGT CCAGCTTCTCCACGGAACAA EcoRV 3 5,592,985
26 GT03009 TCTACCTGCCCAGGAAGCAA TTTCAGGCAAAGCAAAGCAG EcoRV 3 9,205,476
27 GTS03002 AGCATCACGATGATTCTTTTGC TTATTGTGCCCTAGACGAATGA SacI 3 11,547,474
28 GTS03004 TAGGGTTGGCTCTCAATCTCTC ACACCTACAAGGTTGTGACTGC EcoRI 3 13,593,894
29 GTS03008 ATCGCACGGAAGGATAACTG GTTGCTCTAGAACCTCGCAATC SacII 3 19,416,808
30 GTS03009 ATTATCTCACGACCTGGGACTG AGCTAGCTAGGGTTTGTGTTGC EcoRI 3 20,345,341
31 GTS03010 TGAGGAAGAGGAGGAGATTGAG CTGCTTCATTGTGTCTGAGGAC SacI 3 21,733,967
32 GT03021 GAATCTGATTGCCCCCTAGC GGACCCAAATTGAAAGGCAAT EcoRV 3 25,459,877
33 GT03024 TTGGAAGCTTGAAACGATGG TCGCAAGTTCGAAACCCACT EcoRI 3 28,056,489
34 GT03025 TGCGAATACAATTCGGACTGC CGTGCAGTACCCTCCAAACC EcoRV 3 29,184,626
35 GT03030 AATTAAACTCCGCGCCTGGT AAACCATGCATCCGATAGGG HindIII 3 36,245,706
36 GT04004 TGGCGACAAACAGAGCACAC CCTTCTTGCGCAGGACCTC EcoRI 4 4,875,618
37 GT04005 CCAGCCAAACTCAGTTGTCCA CAGACCATACCTCCTTCAGAGCA EcoRI 4 5,924,429
38 GTS041424 ACTGGGAAGAAGGCAAGACA GTGAGCATCCCACTCTCCAT EcoRI 4 6,623,655
39 GTS041472 CCTGAAAGGGCTCAAAAACA TCTTGCAGGTGACAAATCCA PshBI 4 6,914,922
40 GTS041749 TGAACTATTGCGGTTGACCA TGCGGATTGTGTAAGACAGG EcoRV 4 8,481,173
41 GT04014 TGGATCGCTTGATGTGGTTG ATGCCTCAAATACCGCAGCA EcoRI 4 14,685,347
42 GTS043975 TTCGAGGGTTGGTGACTGAT AGAGGGTCCATGGTTTTGTG EcoRV 4 16,607,468
43 GT04020 TCCTTGCAAACGCAAAGTCA GCAACAAAACATTGCGAAGG EcoRI 4 21,083,274
44 GT05001 TTGCAGCTGAAGAAAATGTG TTTGGCACATGAAAATGATG EcoRI 5 602,152
45 GT05003 TGTTGCAGAGTAGCAAACCA TATAAGCTGCAGCCTGGAAT EcoRV 5 2,107,453
46 GTC05003 CCGAAGAGCGACTCCTAGTG AGAGATGCACAGAGGGAGGA PshBI 5 6,043,943
47 GT05013 GAGCATAAGGGGTTTCCCTA TAATGCTGGTGTGGACTCAA EcoRI 5 15,946,504
48 GTC05008 TAATTGCCTCCCAAACAACC TGTCATGCAAAGCAAAAAGG PshBI 5 18,530,587
49 GT05015 CGACGACTATATGGCGGATA AACTCCAATCGGTCATACCA EcoRI 5 20,548,891
50 GT05016 CCGTTTTGTCAGAAACCTGT TAAATCGCGAGACGAATCTT EcoRV 5 22,331,098
51 GTC05010 AGGTGGTGGTGGTTCCATT TCACTGCCGATTTCTCACTG PvuII 5 25,145,098
52 GT05024 TGATCCCCAATGAAAGCTAC TCTTGTACTCCCTTCGTTTCA EcoRI 5 29,110,069
53 GT06013 TCCAACAGCAACAATCTTCA TCTCAAGCTCAATTCCGAGT EcoRI 6 24,403,695
54 GT06015 AGAGCAGAGCTCGTGTTAGTG TGGTGATTTTTGGATGGATT EcoRI 6 26,356,825
55 GT06016 GCGAAGATGCTAGTGGAAAA GTTGCAAAGGATCTGGAAAA EcoRV 6 27,324,979
56 GT06017 GATTCGACGTCTGAAAAGGA ATATGCATGGTTCACTGCAA EcoRI 6 29,168,500
57 GT06019 TGCTCAGCAGAGTCTTGCTAT GCTTGAGACTGGTTCGTGTT SacII 6 31,015,468
58 GT07002 CAAAAGTAACCAATGGCAAGA GCACATTTGAACTGCATCTG EcoRV 7 5,619,862
59 GT07008 TGGAATCAAAGTGAAGGTCAG TGGCAATCTAAAGAGGAGGA EcoRI 7 17,797,502
60 GT07010 TATGGGGACTCCAATAAGCA TACGGTCTCGTCGGTATCAT EcoRV 7 19,489,327
61 GTS07491 AAGGCCAAATGGAGGTTAGC CTGGGAGTCCAGTCATCACA BglII 7 21,220,123
62 GT07013 ATTCAGCCCCTTCTGGATAC ACCTAGGGCAGTCCTTGATT EcoRI 7 22,517,510
63 GTS07984 GCCGCTTTCTAAAAGGATTT GACCGAAATTTTGGCTCAAT EcoRV 7 23,387,924
64 GTS07117 AGCCTGCCACTCTTCCTGTA CGAGGACATGGCATCCTATT EcoRV 7 27,812,959
65 GT07018 TTCAACCTTAGGCACTCGTC TGAGATTCTCCTTGCGTGTT EcoRV 7 28,923,650
66 GTS08007 GATCAGTCGTCCTCGTCCTC GCATTGAGCACATCCATGAC HindIII 8 1,150,395
67 GTS08142 ATGCTCCAATTGAGCTGACC TACTCATGCAGGAACGCATC EcoRI 8 6,123,634
68 GTS08218 AAATGGTGTTTTTCCCGTTG AGGTCGTGGCATGATTTAGC PshBI 8 8,787,812
69 GTS08298 CGATCGGCTCTTACATGACA AGTGGATTTGCAGGCAGCTA EcoRI 8 27,252,203
70 GTS08328 GAGTTACAAACCGCCTTTGC TTGCTAGGGGTCAAGGAAAA BglII 8 28,243,051
71 GTS09004 AGCACCCAGAGCAACAGTTT CCCACTAATTAGCCCATTGC EcoRV 9 620,373
72 GTS09063 GCTTGGATGACAAGCGTTCT AGATCGGCTTCCGAAAGAGT BglII 9 9,835,918
73 GTS09106 ACGCGACTGCTAAGGTTCAT CTCGATCGGTCCCTTTCAT EcoRV 9 11,310,635
74 GTS09188 AGTGGTAGATCCCGTTGAGC AACCGGAGACCTTCAGTGTG EcoRV 9 13,099,244
75 GTS09302 GCATGCATCCGTTGAATAAA GAGATGCTCGTCAGGGACAC HindIII 9 16,712,838
76 GTS09305 ATGGCTTTCGGGCATAGTAA GGATGGCCTATTCAGCAAAA EcoRV 9 21,161,450
77 GTS10005 TCTCAGCCACTCGTTGAATG TCTGGATGGGCTAAATGAGG EcoRV 10 721,349
78 GTS10029 CCTCCCTTCAGTTTGTGAGC CATTGGATCCCCAAAGAAAA EcoRI 10 3,044,726
79 GTS10116 CCCCCTAAGATGGTTTTTGC TGAGAAGTGAAGTGTGAGTGTGG EcoRV 10 4,577,975
80 GTS10199 AGCCTGAGAAGCAGATTGGA TGCAATGACGTGATGGTACA EcoRI 10 6,149,421
81 GTS10204 TTTCACACGGGACATTCAAA TCGACCTTGGACTTCTAGGC StuI 10 9,961,760
82 GTS10214 TAGGATCTCGCCGAACATCT ATGCACCTCCCCAACATATC EcoRV 10 11,290,285
83 GTS10276 TGGTCTTTGAAGCATTCTGG TTCAATATTGGCCGTAAACC EcoRV 10 13,637,268
84 GTS11014 CTTCGCTTCATTTCGAGTTTG CATTTCTGCAGTCGCATTTG EcoRV 11 353,397
85 GTS11016 CGCACGAGATGCTGTCTAAC GAAAAACCCTCTGCCCTCTT NruI 11 1,389,204
86 GTS11086 TGCTGGTGTGCTATTTGAGC CGTGGCCATCCAAAAGTAAT EcoRI 11 17,330,622
87 GTS11088 CGCCTAGGTGCAGAAAAGAG ACACCATGAGCCTCCTCACT BamHI 11 20,838,960
88 GTS11330 TGTCATTGGTTGCCCTGTAA AGCATGGAGGTGGTGCTAAC EcoRV 11 26,534,991
89 GTS11350 AGGAATATGAATCGCGTGTG TTTGGGAGCTATGGCCTATG EcoRI 11 27,197,820
90 GTS120003 GCGTTTTTCCTCCAATTCAA ACCGACAACTTTTGCCCTTT PvuII 12 3,007,338
91 GTS120032 AACGAAGAGGCCAATGGATA GTTGACCATGGGTGCTCACT PvuII 12 3,807,192
92 GTS120139 TGCTTTCAGGAGAGCAGGAT CCACCAGCATCACTCTGCTA EcoRI 12 9,263,934
93 GTS120457 GCATGAGAGACAGCGGAGTT GTGATTTGCCATGCCTTTTT EcoRV 12 13,674,598
94 GTS120652 TGGGCACAACTACAAAGGTG CATTCCCATGTTCCACATCA EcoRI 12 17,438,893
95 GTS120760 CCCAGCCACAAGAAATGAAT TTTTCCTTTTCCCTGCTGTG EcoRI 12 19,070,762
96 GTS120842 AAGCCATCATGTTCCATTCC TTTTGGTACGTCATCCGTGT EcoRI 12 21,522,146
97 GTS120941 CAGGTGGTCCTTTTCAGCAT TAGTGTCATGTGCCCTGAGC EcoRI 12 22,565,939
98 GTS120989 CCTTTGGAGGGCTTGACATA TACCCACTGGAAACGGAAGA EcoRV 12 23,622,982
99 GTS121030 CGTTGCTCCGTTCATCTTCT GCTAGCCCCTTTTCATCTCC EcoRV 12 26,276,873

Primer sequences of Bph3IND and BPH26IND markers.

marker name      forward primer      reverse primer
Bph3IND GCTTCACCTGTGTCCTGTT GTGTATGAGGGTTGTTCTGGAT
BPH26IND GGGCAGCTAAGCGCATGGTT GGGTGATAAATCGAACAGCAGAGG

Identification of QTLs for BPH resistance

QTL name chromosome location (cM) QTL interval* (cM) LOD additive effect dominance effect R2
 qBPH4 4 0 0-6.6 7.22 0.94 -0.20 0.138
 qBPH12 12 51.4 40.8-54.4 6.64 0.87 0.55 0.131

*interval at 99% probability

Table 1 The list of primers for amplification of Bph3 and BPH26 genes of Gayabyeo

* Bph3 is composed of three genes that are OsLecRK1, OsLecRK2, and OsLecRK3.

** Sequencing of BPH26 is done on its three exons according to Tamura et al. (2014).

Table 2 List of developed CAPS markers
Table 3 Primer sequences of Bph3IND and BPH26IND markers.
Table 4 Identification of QTLs for BPH resistance

interval at 99% probability

Table 1

Table 2

Table 3

Table 4

ABOUT

BROWSE ARTICLES

FOR CONTRIBUTORS

EDITORIAL POLICIES

© The Korean Society of Breeding Science.