A genetic linkage map of allo-octoploid strawberry (Fragaria × ananassa Duch.) using SNP markers

Ye Rin Lee; Jundae Lee

doi:10.9787/KJBS.2017.49.3.119

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SNP 분자표지를 이용한 8배체 딸기( × Duch.) 유전자지도 작성

A genetic linkage map of allo-octoploid strawberry (Fragaria × ananassa Duch.) using SNP markers

Ye Rin Lee, Jundae Lee^*

Korean Journal of Breeding Science 2017;49(3):119-128.
Published online: August 31, 2017

DOI: https://doi.org/10.9787/KJBS.2017.49.3.119

Department of Horticulture, Chonbuk National University, Jeonju 54896, Korea

*Corresponding Author : (ajfall@jbnu.ac.kr, +82-63-270-2560, +82-63-270-2581)

• Received: April 6, 2017 • Accepted: June 9, 2017

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.

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적요
서 언
재료 및 방법
결과 및 고찰
적 요
사 사
References

적요

An allo-octoploid strawberry (Fragaria × ananassa Duch.) is one of the most important vegetable crops in Korea. However, there were few genomic researches of strawberry due to polyploidy and complexity of its genome. In this study, we aimed to construct a genetic linkage map of strawberry using single nucleotide polymorphism (SNP) markers that were developed through a next-generation sequencing (NGS) analysis. Two strawberry varieties, ‘Sulhyang’ and ‘Senga-sengana’, were used as a maternal and a paternal parent, respectively, and their F1 generation consisting of 94 individuals was used for construction of a genetic linkage map. A total of 19.0 Gbp (‘Sulhyang’) and 21.8 Gbp (‘Senga-sengana’) of genomic sequences were obtained through NGS analysis. Subsequently, approximately 87,000 SNPs were identified and 1,154 primer sets for high-resolution melting (HRM) analysis were designed through bioinformatic analysis. In result, a total of 224 polymorphic HRM markers were developed and 205 markers were mapped on the genetic linkage map of strawberry, which total length was 800.8 cM and the number of linkage groups were 30. This SNP-based genetic linkage map and the 224 SNP markers will be very helpful for the genomic and genetic researches of allo-octoploid strawberry.

서 언

현재 국내에서 재배되고 있는 딸기(Fragaria × ananassa Duch.)는 8배체(2n=8x=56)로 F. chiloensis와 F. virginiana가 자연적으로 교배되어 만들어진 종으로 알려져 있다(Darrow 1966). 딸기의 배수체는 2배체(2n=2x=14), 4배체(2n=4x=28), 6배체(2n=6x=42) 및 8배체(2n=8x=56) 등 다양하게 분포하고 있다(Folta & Davis 2006). 그 중 2배체 종인 F. vesca에서 유전체 연구가 가장 활발하게 진행되었다(Shulaev et al. 2011, Folta 2013). 또한 딸기의 유전자지도도 F. vesca에서 simple sequence repeat (SSR) 분자표지를 이용하여 제일 먼저 작성되 었으며(Sargent et al. 2004, Monfort et al. 2006), 2011년에는 F. vesca의 유전체 전체 염기서열이 공개되기도 하였다(Shulaev et al. 2011). 결과적으로 F. vesca의 이러한 연구들은 다른 Fragaria 속에 속한 종들의 유전자지도 작성 및 유전체 분석 연구에 큰 기여를 하였으며(Rousseau-Gueutin et al. 2008, Sargent et al. 2009 & 2012, Isobe et al. 2013, Hirakawa et al. 2014), 이와 동시에 장미과(Rosaceae) 내의 비교 유전체 연구의 기초자료로 이용되기도 하였다(Vilanova et al. 2008, Jung et al. 2012).

반면에 8배체 종인 F. × ananassa의 경우, 영양번식을 하는 이형접합형 작물이기 때문에 경제적으로 중요한 작물임에도 불구하고 오랫동안 유전자지도 및 유전체 연구가 진행되지 못 하였으나, 최근에는 그 중요성이 더욱 부각되어 다양한 연구가 수행되고 있다. F. × ananassa의 최초 연관 지도는 amplified fragment length polymorphism (AFLP) 분자표지를 이용하여 작성되었고(Lerceteau-Köhler et al. 2003), 그 후에는 주로 SSR 분자표지를 이용하여 유전자지도가 만들어졌다(Rousseau-Gueutin et al. 2008, Sargent et al. 2009 & 2012, Isobe et al. 2013, Van Dijk et al. 2014). 또한 F. vesca의 유전체 정보를 이용하여 F. × ananassa의 유전체 분석 연구가 수행되기도 하였다 (Hirakawa et al. 2014). 특히 Isobe et al. (2013)은 1,856개의 SSR 분자표지를 이용하여 총 2364.1 cM의 연관거리를 가진 28개의 연관군으로 구성된 고밀도 유전자지도를 작성하였다. 그러나 SSR 분자표지를 분석하기 위해서는 폴리아크릴아마이 드 젤 전기영동을 수행해야 하기 때문에 많은 노동력과 비용이 소모되 고 또한 대량 유전자 분석 시스템(high-throughput genotyping system)으로 개발하는 것도 쉽지 않다. 따라서 SNP 분자표지 개발이 절실히 요구되고 있는 상황이다. Bassil et al. (2015)과 Sargent et al. (2016)은 8배체 딸기의 SNP 유전자지도를 보고한 바 있는데, 이는 90K Axiom^® SNP array를 이용한 것이다. 대량 유전자 분석 시스템(high-throughput genotyping system) 은 대량의 분자표지를 한 번에 분석할 수 있지만 개체당 분석비용 이 비싸기 때문에 현재 상황에서 실제 육종 프로그램에 활용되기 위해서는 한 개씩 분석할 수 있는(flexible) SNP 분자표지가 필요하다.

따라서 본 연구에서는 양친의 염기서열을 분석하여 대량의 SNP를 탐색하고, 이를 이용하여 SNP 분자표지를 개발하고, 개발된 SNP 분자표지를 이용하여 8배체인 F. × ananassa의 유전자 연관 지도를 작성하고자 하였다.

재료 및 방법

식물 재료

식물 재료는 국내 품종이고 역병 이병성인 ‘설향’을 모본으로, 독일 품종이고 역병 저항성인 ‘생가생가나’를 부본으로 사용하 였고(Eikemo & Stensvand 2015), 이를 교잡한 F1 분리집단 94개체를 이용하였다.

식물 DNA 추출

교배 모본으로 사용한 양친과 94개체의 F₁ 식물체로부터 각각 DNA를 추출하였다. 1.5 mL microcentrifuge tube에 2-3매의 딸기 신초, 5 mm stainless steel beads 2개, DNA extraction buffer (20 mM Tris-HCl, pH 7.5; 250 mM NaCl; 25 mM EDTA; 0.5% SDS) 700 μL, 1 g PVP (polyvinylpirrolidone), 및 12.5 μL beta-mercaptoethanol을 넣은 후 Tissue Lyser II (Qiagen, Germany)를 이용하여 3분간 마쇄하였다. 그 후 70°C 의 Lab Armor beads bath (Thermo Fisher Scientific Inc., USA)에 10분간 열처리를 한 후 원심분리기(Hanil Co., Korea) 를 이용하여 4°C, 13,000 rpm에서 10분간 원심분리하였다. 상층 액 600 μL을 새로운 1.5 mL tube에 옮긴 후 동량의 chloroform: isoamyl alcohol (24:1)을 넣고 4°C, 13,000 rpm에서 10분간 원심분리하였다. 이 후 다시 상층액 500 μL을 새로운 1.5 mL tube에 옮긴 후 동량의 isopropanol을 넣고 -20°C에서 10분간 보관한 후 4°C, 13,000 rpm에서 10분간 원심분리하여 DNA pellet을 얻었다. 그 다음 70% ethanol을 700 μL 넣고 4°C, 13,000 rpm에 1분간 원심분리를 하는 방식으로 2번의 세척을 하였다. 마지막으로 0.1 μL RNase와 100 μL 멸균수를 섞은 용액을 DNA pellet이 들어있는 1.5 mL tube에 넣어 용해시킨 후 BioDrop Lite (Biochrom, England)를 이용하여 DNA 농도 를 측정하였다. 추출된 DNA는 30 ng·μL^-1 농도로 맞추어 사용 하였다.

차세대 염기서열 분석 및 SNP 탐색

대량의 SNP를 탐색하기 위하여 추출된 ‘설향’과 ‘생가생가나’ 의 DNA 1 μg씩을 이용하여 차세대 염기서열 분석(next generation sequencing, NGS)을 수행하였다. 생물정보학회사인 씨더스 (Daejeon, Korea)에 의뢰하여 NGS resequencing 분석과 생물정 보분석을 수행하였다. 딸기 표준유전체 정보는 Strawberry GARDEN 홈페이지(http://sstrawberry-garden.kazusa.or.jb/)에 공개된 Fragaria × ananassa (FANhybrid_r1.2)을 이용하였다 (Hirakawa et al. 2014). 딸기 표준유전체의 총 염기서열 길이는 720 Mbp였으며, DNA 단편조각인 387,847개의 scaffold와 238,119개의 contig로 구성되어 있었다.

대량 분석된 염기서열은 전처리, 표준유전체에 정렬, SNP 탐색, non-paralogous SNP 선발, HRM 프라이머 디자인의 과정 을 거쳐 분석되었다. 전처리 과정은 solexaQA (v.1.13) package 의 DynamicTrim과 LengthSort 프로그램을 사용하여 분석하였 고(Cox et al. 2010), 전처리 과정을 통과한 cleaned reads를 BWA (0.6.1-r104) 프로그램을 사용하여 표준유전체에 mapping하였다(Li & Durbin 2009). 그 결과 생성된 BAM format의 파일을 SAMtools (0.1.16) 프로그램을 사용하여 raw SNP를 탐색하였고 동시에 consensus sequence를 추출하였다 (Li et al. 2009). 그 다음 ‘설향’과 ‘생가생가나’의 SNP를 비교 분석하기 위해 SEEDERS in-house script를 이용하여 샘플간 통합 SNP matrix를 작성하였다(Kim et al. 2014). 마지막으로 다형성 SNP의 flanking sequence (400 bp)를 추출하고, 딸기 표준유전체에 BLASTN (2.2.26+)을 수행하여 서열 비교를 통 해 딸기 표준유전체에 한 번만 나타나는 SNP를 선발하였고, target SNP를 증폭할 수 있는 high-resolution melting (HRM) 분석용 프라이머를 디자인하였다(Table 1).

Table 1

List of HRM primer sets used in this study.

Table 1
Marker name	LG	Position (cM)	Forward primer	Reverse primer	Fragaria vesca genome

					Chromosome	Position (bp)

FAN-SCF420	1	0.0	GGAATACCGAGACTCGTCACT	ACGACAATCCTATTCAATCCACTG	6	30689884 - 30690283
FAN-SCF813	1	9.0	TGAATAAAGGGTTTTCAGTTTTGA	TCAGACATCTCTCAAGGTCGT	6	30647220 - 30647488
FAN-SCF384	1	10.2	ACCAACAACTCAAACTTCAAAAGT	GGCATTTTGTTTTATGACGAGACT	6	31592825 - 31593037
FAN-SCF132	1	11.4	AGTGGAGGACATATTCAAGTGTCT	GTTGCGGTGCTTAAGAGTGC	6	31401845 - 31402250
FAN-SCF418	1	12.8	TCGAAATTCACACGAAGAAGA	TTGGACTATACTCTGCAATACTG	6	31347505 - 31347862
FAN-SCF892	1	15.4	TGGGCCACTCTGTCTATGTT	TCGGCTTGTCCATCCTTTCC	6	31458344 - 31458618
FAN-CON1139	1	21.4	TGCAGGGGACTCTGAGATCA	TGGAAGTCCAGGAGTACTTGG	6	28464183 - 28464582
FAN-SCF833	1	38.6	TGAAGTGCTGCAATCCTGGT	AACGGTGGGCTAGATTTGGG	6	25589098 - 25589497
FAN-SCF174	1	66.7	TGTTGTAGTGAGAGCCGTCG	ACCAAAGGGCGATGATCCTC	6	15978363 - 15978658
FAN-SCF715	1	71.4	TGAAGGAAGAGGAGGAAGTGT	TGGTTTAGGGTGTTTTAGGGGT	6	14511815 - 14512028
FAN-CON1046	1	78.9	CTAGCCTGGTCAACCGACTC	TCCCATTAGTGTGGTTTAGCGA	6	10237152 - 10237439
FAN-SCF338	1	102.0	TGCATACATTTTCTGTGCGA	CCAGGTAACGATAAGGGCCC	6	7114971 - 7115361
FAN-SCF735	1	103.8	TGCGACAGGATAATCATGACGT	ACCCCTACTCCATCATCCCC	6	7158469 - 7158868
FAN-SCF858	1	113.4	AGAATCTAGACTGATGGATCGT	ACCGACGTAATAACAACCACCA	6	4581492 - 4581890
FAN-SCF33	2	0.0	CTACTGCCCACAACTCGGTG	CCAATGGCTACAATCTGCGC	7	16926268 - 16926609
FAN-SCF607	2	68.5	CTCGTGAAATTAGCATCCGTCA	TCTCTATGCACACTGACAACCA	4	24082358 - 24082757
FAN-SCF377	2	70.8	GCTGATGAAACATAGTAATTCCGA	TGCGGACAAAATGGATATTTTCT	4	26910570 - 26910801
FAN-SCF468	2	72.0	ACCAAAATCATAAGCAAAACCAGA	TCAGAAGGAAGCTTTCTAGAGCA	4	26624055 - 26624454
FAN-SCF399	2	72.0	ACCTCTCAAATGTGGTTCTCA	CCCTCTAGGACCTTCTGATGC	4	26707960 - 26708303
FAN-SCF405	2	72.0	TCCACAAGAACCTAAAAGCAT	GGTGAAGGTAAATGAAAAATCTGT	4	26145596 - 26145936
FAN-SCF719	2	72.0	AGCCTCCCAGCCTAACAGTA	CATGATTCAATGACGCGGCC	4	26613015 - 26613412
FAN-SCF39	2	73.1	CATGGCAGGCTAGCTTCGAT	TGGACCGACTTCCTCAACAG	4	26359016 - 26359415
FAN-SCF52	2	74.7	TGCTCTTTCGAAATGCCAGTG	TCACACACTGCTTAGCAACTGA	4	26652718 - 26653117
FAN-SCF956	2	77.0	ACTCTTAGCTAAATGGTTCTTGCT	ACCCTATATTGCAAGAACACCT	4	24693600 - 24693962
FAN-SCF824	2	79.2	TTCCCTATATATGTTTCCCATGCA	ATGCGCACCACCGAGCTTAG	4	24568435 - 24568834
FAN-SCF309	2	80.8	CCAATAAAACTTGCGGCGGT	TGGTGGGTGCAATTTGATTG	4	24851133 - 24851532
FAN-SCF637	2	81.9	ACTAGGAGGAAGGGGCCTTT	ACACCAAGATCAGCATCATCA	4	26476579 - 26476858
FAN-SCF578	2	86.0	GCTATTCAGCGGGGTAAGCA	GCTTAGTGTCAAAAATGCAGCC	4	25972133 - 25972532
FAN-CON1146	2	88.7	AGTCTCCAAAGCCTGAAACCA	CCAGGTATTCGGTCAGTTGCT	4	25186628 - 25187027
FAN-SCF6	2	89.2	TCGCAAATGGCTTTTTAGTTGC	AGACTGTATTTGTCTGAGACGA	4	25176196 - 25176558
FAN-SCF131	2	89.8	ACAATCAAACGTGAGGGGCT	CCAACGATGAATGTGAAGAACAGA	4	25687752 - 25688153
FAN-SCF909	2	94.5	ACCCCTAACTAAGTCATCACCG	CCTTGTTGGGTCTGGGCTAA	6	21985116 - 21985169
FAN-SCF945	2	96.7	TGCTGCACATTGTTCAGTCT	TGTTTATTCAGTGTTCATCAGATG	4	23040342 - 23040504
FAN-SCF595	2	98.5	TGGTGCAGGATGAGAATGAGG	CACCTGGTGACGGTCATTCA	4	22850917 - 22851309
FAN-SCF345	2	102.6	AAGGGGACGGTACAGCTACT	GGTGCCATCCATTGAGGCTA	4	22292764 - 22293102
FAN-SCF163	2	103.9	GGTTCGCTATGGCTAGGTCC	AGCTCAAGTTGTGGGTTCCA	4	22363717 - 22364056
FAN-SCF231	2	104.5	ACTGCTTTCCTCTGTGGTTT	AGAGGCATACAACACTGAGTCT	4	22008076 - 22008475
FAN-SCF294	2	106.8	GTGCCTTCAAGGAGCTGGAT	CCCTGCCATAAATTGTTAAATGCA	4	22202764 - 22203163
FAN-SCF402	2	109.1	GCACGCCTCCTCAGTTACAT	CGAGCTGACGTTTAAGTCCCA	4	22640567 - 22640947
FAN-SCF108	3	0.0	AGGGTGTGGAATATCATCACTCA	ATGGGCCTTTGTACCGGTTT	6	21857853 - 21858252
FAN-SCF389	3	22.4	TCTAGCGCAATCTGCACCAA	TCAATGAAACGTACGGCGGT	6	21995951 - 21996362
FAN-SCF166	3	26.1	TACACCGACAGGGAGGTTGA	TTGATCCGAGCGAGCTTACC	6	22152785 - 22153184
FAN-SCF59	3	26.1	GGCTCTGACTTCGGCTTCTT	TCTGCTGGCATTTTGTGTGC	6	22245209 - 22245570
FAN-SCF51	3	26.1	CAGGAGGGACTGGAAAGACA	TGTGTTGTCATCCCACTAAGCA	6	22330843 - 22331242
FAN-SCF135	3	26.7	TCGTTGGAACCTCAGGTTGG	AGGGATATTGATGATATTGTTGCA	6	21563283 - 21563682
FAN-SCF185	3	27.2	GTGCTACCGCTGAGGACAAT	CTAACATCCTCCTGGGCGTG	6	21786636 - 21787035
FAN-SCF105	3	27.2	TTTCGCTTCCAAGTCACCAAC	AGTTTGTCAAAATCTCACCTCC	6	21410739 - 21411138
FAN-SCF215	3	27.8	CGTCGTAGTGGATGATCCTT	TGACAAATGACAACCCATGTT	6	22851692 - 22852092
FAN-SCF58	3	28.3	TGGGGTAAGTGCATTTGGTCA	CCAAAGCCACTCCTCTTCGT	6	23743188 - 23743587
FAN-SCF83	3	28.3	ATGGGCTCCAGCTCTAGTCA	AGAGAGAGTTCAAAAGAGGGGT	6	23220621 - 23221030
FAN-SCF115	3	28.3	GGTGGACATGAATATGAGAAAGCA	GGAAAGACCCCTACAAAAACCC	6	23209827 - 23210225
FAN-SCF173	3	28.9	ACCAGACTACCACTGTGCCA	AAGCAGGGTCGACAACTACC	6	23136810 - 23137047
FAN-SCF388	3	30.7	ACCCTGCCTCCTGTTCAAAC	CACGGCTATACCCAAAGAGCT	6	24286158 - 24286551
FAN-SCF437	3	33.2	TTCTTGCAGGCAGACCAGTC	GGTGCACTGAGGTGGTTTCT	6	22909018 - 22909416
FAN-SCF238	3	33.8	AGCTGTCTAAAAAGCTCTTCGGA	TCGCAACAGAAGTGACTCCC	6	21833742 - 21834148
FAN-SCF450	3	33.8	CCGAAACATGGTCGATCCGT	CCCAACCTTGCTGCTATTGC	6	22164873 - 22165272
FAN-CON1153	3	34.4	CCCATACGCGTGGCTTCAAT	ACTTATCCTCTTCTTTGATGTGGG	6	21433559 - 21433813
FAN-SCF438	3	43.0	TTCCCAGATCACTCTGCTGA	GGGTTGTTAAGGTATAGTACTTGG	6	13401292 - 13401547
FAN-SCF436	3	47.2	ACCAATTTATCGAAACTTCACCGA	TCGATCAAAGACTTGCTATCGA	6	14629000 - 14629399
FAN-SCF246	3	48.3	TGAGGAGTAGGAGCTGTCCA	TTAGCTAAGCAGGGTTGGCG	6	10601014 - 10601413
FAN-SCF165	3	50.1	GCACAAAATATTGCTGATTAGGCA	TGCAGTTTCTTATATGTTGTACGG	6	14765076 - 14765398
FAN-SCF79	3	59.0	TGTTGTTTCTTCCTCCTTGGT	AGTTGGTAAAAGTCTAGGTATGGA	6	11024227 - 11024401
FAN-SCF233	3	60.1	CCAACTCAAATGTGAAGTTGTGGA	TCCGCCCAGACTCAAAACTT	6	9124886 - 9125097
FAN-CON1070	3	65.5	GCAAGCTTTGGCACTCATCG	TCCCTCAGGAGATGGTTGGT	6	9045076 - 9045396
FAN-SCF443	4	0.0	TCACAACGCTTGTCAAAGGC	ACCAGAAAGAGTATCCTCGTCT	-	-
FAN-SCF172	4	3.5	TTGAGGCTCATTGACAACGT	TGAGGGAGAGAGATAGAGGTGC	6	3493659 - 3493907
FAN-SCF316	4	6.5	ACAGCCTCAATCAGCTGCTT	TAAGTTTGGGCGATGATCCT	6	2504998 - 2505390
FAN-SCF769	4	30.7	CCGAAGAGATGAGCAACGGT	GGGACTTGATCATTTGCCGG	6	9394367 - 9394762
FAN-SCF656	4	32.5	ACATATCGCTGCTGGACCAC	AGCTAGCAACTTCCGACCAA	6	10822978 - 10823285
FAN-SCF119	4	34.6	CCATCATTTTACAAACGCTACACA	ACGTGATCTTGCAACGGGAA	6	11132414 - 11132696
FAN-SCF569	4	35.3	TGACTTTTCGTCCATGTTGTCA	GACCGAAACGTATCCATAAATAAA	6	10946444 - 10946849
FAN-SCF570	4	37.7	ACGAAAGTGGAGGTGCTCTG	TGAGCGAGATGAAAACGTCG	3	5315532 - 5315808

FAN-SCF273	4	43.8	TCTCAAACGTCTTGACGCCT	TGAGTGCTATTGTGGCGACT	6	18744174 - 18744235
FAN-CON1062	4	43.8	ACACCACGATATGCAACCACT	ATTTCAAATGGGCCGGGTCA	6	14238062 - 14238401
FAN-SCF183	4	44.3	ACCATCGACGATGAGGAAGC	CGGGGATGCTGGTCATACTT	3	31695069 - 31695130
FAN-SCF69	4	48.8	AGCGGTCCATTGACTCAACA	TGTTGTAAAGCTTGTCGGGA	6	14700987 - 14701320
FAN-CON1071	5	0.0	TTGAGGTTTAAGCCGAGATGACT	GTACTAACAGAGGCGGCGTT	6	27458588 - 27458625
FAN-CON1087	5	6.2	TGCAGATGTTGACTCTGCGT	TTTTTACGTCCGCCGACTGA	4	32415681 - 32415732
FAN-SCF290	5	7.3	TTCACTCAAAAATTCGTGTGTT	ACTTCTTACACTGACAAAACTCGA	4	32269269 - 32269656
FAN-SCF168	5	35.6	TCAACAGAGAAGATGCTTGGT	ACACTCCCTTTTCCTAGCACA	6	9735998 - 9736029
FAN-CON1123	5	37.6	TCGTCACATCTACAATTGGCTCT	GCGATGGTTCTAAGGTGTTCG	4	23313914 - 23314024
FAN-SCF965	5	39.3	TGTGGAGAAGTTCTAAGAATCCCG	CGATCAAAGATGAAACCTAATTCC	6	1903611 - 1903648
FAN-CON1056	5	42.8	AGTGATTGGGCTGCATCAAA	GCTAGGTACAGTTGCCCTGG	4	21935175 - 21935548
FAN-CON1143	5	47.0	TGGTAGCCTTCCCAATGCAG	ATAAGCCAACCCCCGAGTTG	5	11569758 - 11570157
FAN-CON1128	6	0.0	TCGACATAGTGAACGGTGGC	CAAATGGATTTGCGGCACCA	7	423374 - 423768
FAN-CON1053	6	1.1	GCTGCAACAAAATTGGCTTCC	AGGAGACTCTAAGGTATTGATCGA	7	344631 - 344903
FAN-CON1059	6	2.3	CCTAGCTCGGCAATCAGAGG	TGCAGTCAGGAGCAGTTGAG	6	30104705 - 30104918
FAN-CON1097	6	15.3	ACGTTGACGCGTAATTTGGA	TGCAGTTGGTGAATGTTTGA	7	6499804 - 6500187
FAN-SCF474	6	19.3	TCGTGGGCTTAAGTTCTCAAGA	CTCGCCAAGAGTATTCTCCA	-	-
FAN-CON1122	6	21.0	AGGACTTCTTTGATAGTACCGACA	TCTTCCCAACCCACAAGCAA	7	7598690 - 7599115
FAN-SCF145	6	23.4	GCAATTTCTAGAGAGGCAAGCA	TGCTTTTAAAGACAGAGAAGACA	7	5750781 - 5750840
FAN-CON1073	6	33.3	ACCCGTGAAACAATAGGGAAC	AAAGCGATATACATAACATGCACC	7	11248187 - 11248367
FAN-SCF961	6	34.6	TGTGCGAATTTAATTTCCACACA	GAGGAAGCATGAGCATCGGA	1	2921841 - 2921960
FAN-SCF139	6	41.1	TGCAACTCAACCAAGCCAGA	AGTTCACACTTTGTAGCCACTGA	7	12594627 - 12594928
FAN-SCF730	7	0.0	ACCGGGTAACTTTGTTGGCA	ACTCACCAATATCCGATGTGGT	2	10795356 - 10795437
FAN-CON1134	7	1.7	ATCTGAGGTTTTAATTCTCATGC	TGGTATTGTACAAGGAAAGAGAGG	7	2111414 - 2111539
FAN-CON1145	7	2.8	TGGAAGCACTAGCCTTTGTCA	GCAAAAGGAAAGAAATAACGTGGG	5	8070408 - 8070518
FAN-CON1115	7	23.5	TCATTTCCTGGCAGCTACTGA	TCAGAAACCAAACTAGCCAAT	5	11182526 - 11182760
FAN-SCF756	7	32.7	GCTACTGCATTTTAGCACTTTGC	TTGATAAACCAGTCTTATAACGCG	5	24691171 - 24691218
FAN-CON1066	8	0.0	CCTTTTGGGCCCATGACTTG	GGTGTGATTAATCGGCCCGA	5	8945836 - 8946223
FAN-SCF378	8	0.0	CAAGAGGCAGCTTCTCTGGT	GACTCCACAAAGCCTGGTCA	5	9274286 - 9274540
FAN-CON1069	8	9.7	TTGTCGTCTTAATCAAATGATTGT	ACAGCTATATCCTCCACATGT	5	5735911 - 5736310
FAN-SCF698	8	12.0	GCCGAAAACCGTGGTTTGAG	ACCGTGCTTGTTGTACTCGT	6	4563039 - 4563335
FAN-SCF27	8	12.5	CTGTGACTCTGTGAGGGCAG	TGCAGACATGAAATGCACCA	5	5272835 - 5273234
FAN-CON1117	8	15.4	GTTTCACCATCCAAGCTGCG	TCAACCCACAACCACCAACT	5	4918889 - 4919285
FAN-SCF227	8	16.0	TCACCGAGAGTAGCCTAGGA	AGATGCCAGGTTACTAGTTACT	5	4858458 - 4858855
FAN-SCF63	8	25.0	GTTCTGATTGCTAGCATCTGCA	TCCTGATGCGCTACTCTTGT	4	32169627 - 32169969
FAN-SCF480	8	27.3	AGGATAACAATCTGTAGTGGGGA	TCACCTTAGGCCTCAGCAAA	5	2964486 - 2964724
FAN-CON1015	8	30.2	AGATTGTTGGACGCCTCTAGTC	TCCTCATCAGATAAGGTGTGCA	5	1708322 - 1708548
FAN-SCF260	8	30.8	TCTCTTGAACACATTGAACCACA	TCTATAATCGGGTTCTTGGGT	5	1188138 - 1188537
FAN-SCF434	9	0.0	ACGTTGGTTTCTAAGGGTGT	CGATCACAAGGTCCTGAGAGA	6	35969226 - 35969306
FAN-SCF279	9	2.2	CCACCTGGGGGTTGAAAGTA	CCGCTTTGCATAACTTTCGGT	2	15447700 - 15447757
FAN-SCF326	9	2.9	TGGTTTTTCGCATTTGTTGGT	GGGGACGCTTGAAGAACACA	6	2118937 - 2119014
FAN-SCF846	9	7.7	TGTCAGACACAAGTTAGCTTTTGA	AATCACGGTCCAACACTCGT	6	35292089 - 35292279
FAN-SCF847	9	18.4	GGAGCTTCTTTATTTTTACTCGCA	TGCAAAATCCGGACAGCAAC	6	32014064 - 32014193
FAN-SCF88	9	26.8	TCATACTTGTTGCTGTAGTCACA	GGGTTCTGTTTGTGGTTGGG	6	17704328 - 17704727
FAN-CON1099	9	30.6	AGGAGAAGGAATTGGATCACA	AGGTTCTTTCTTGGACTGGCA	6	28740651 - 28740822
FAN-SCF369	10	0.0	TCAAGTTCACGTTTTACAGTGTCA	TGCATTCTCACCTTCTCAACT	2	14150750 - 14151150
FAN-CON1089	10	15.0	GGCACAACCTGTTTGAGTTT	AGTCCATTCTGCTCCAGTCT	2	3613082 - 3613418
FAN-CON1105	10	19.8	GCGATGACCCAATTTTCGGG	GCCACATCTGTCACATATGCC	2	10339151 - 10339546
FAN-SCF154	10	20.4	TGCCAGCGAATGAGTAGCAG	ACTGCTTATTTGCTTTGGCCA	2	10167626 - 10168044
FAN-SCF559	10	21.0	GCAAATTGAGTACAGTGACCGA	CCTTGGGTTTGCCTGTAGAA	2	10088172 - 10088571
FAN-SCF1	10	29.6	GATACAGACAGGTTTGTGCGC	TATGGTCTCGCAGAAGCCAC	2	10333513 - 10333912
FAN-SCF605	11	0.0	TGCCCCTCCTCTCGTTTTTC	TGACTTCGAACTTGACCTCCA	6	18216050 - 18216445
FAN-SCF807	11	18.9	TGCAATAAAGCAAAGTTCAAAAAT	AGTTCTTTTGCTTCGTACACT	6	28062711 - 28062898
FAN-CON1016	11	24.9	AGGGAATGGAGGTATAAGAGTT	CCTTTCTGGTAGCAGCCTTACT	6	26052420 - 26052541
FAN-SCF67	12	0.0	AAGGGTGGTTTTAGGGCTGG	CGGGCTTTCACCGATCTCTA	4	7444429 - 7444493
FAN-SCF203	12	1.6	ATTTGCGAACCTGTGACCAC	AGAAAATGATGGCGCTTCAACA	3	8554786 - 8554880
FAN-SCF235	12	6.6	ACCCACTATGGAGTTAGCTCT	TCCAACAAACACCTAGGGCA	6	3571899 - 3572282
FAN-SCF129	12	11.4	TGGCGATACAAGTTCGTAAACA	AGTGAGGCTGATTTAGAGTTTCCA	5	14764214 - 14764422
FAN-SCF2	12	23.9	TCGAGAGGTGTATAAAATGCTCT	TGTTTGTATGCATAACAAGCTGA	4	5849067 - 5849236
FAN-SCF548	13	0.0	CCTGGGGTGTTCAACTAGGC	ATCTGCCGTGTGATGCAAGA	5	10208591 - 10208636
FAN-SCF112	13	5.1	TGTAGGAGACAAAGCGTAGTGA	TGAGTAGCAGCAGACTGTAACA	6	11944125 - 11944534
FAN-SCF34	13	5.7	ATTTGAAACGCGTGGATCGAG	GTCTCCAGCACCACCAGGAA	2	1358158 - 1358239
FAN-SCF84	13	7.3	CTTGAGGCCGACGATCTGTT	AGATTCTGCCATCCCGCATC	3	1653607 - 1653678
FAN-SCF483	13	12.8	GGGCATTGGGATCTCAGCTT	CGTCGCCTAGTGACAAGGAA	6	9565771 - 9566099
FAN-SCF760	13	15.1	CAATGACCCTCACTCACCCC	ACCCTACACCATGATCCTCCA	6	9911049 - 9911186
FAN-SCF263	13	22.8	TGTTTAACGGATCATAAATGCAT	GTCATTCGCGACTCGGTACA	6	12241307 - 12241731
FAN-CON1039	14	0.0	GAGATGAGGATGCAGGCCAA	TGCTCACTCTATCGATGGGA	6	20401011 - 20401336
FAN-CON1142	14	6.8	AGTTCATCCGAAGCTGTGCA	ACTGGATATACCGCGCATGT	6	12579743 - 12579956
FAN-CON1137	14	10.9	TCACCTGGAAATTCAGAGGA	TGCCAAAGGTCGAAAATGCG	6	13012521 - 13012836
FAN-SCF289	14	22.2	ACCCTGAAATGGTTAACGTGGT	GCCATATGCCTTACTCATATCT	6	20589193 - 20589212
FAN-SCF905	15	0.0	GGAGGGCTGTTCTTCAGTGG	TCTTACGCAGTGCAGTCCTG	3	22549713 - 22549953
FAN-CON1076	15	1.2	GGTGCAATCCAATGAGCAGG	ACTCAGGCTCGGTAGACCAT	3	22549227 - 22549626
FAN-SCF362	15	6.1	CTGGATCGCTACCTGCACTC	TTAGGAGGGGTCTTCCGTGA	2	6880598 - 6880965
FAN-SCF66	15	17.8	TCCCAGCGCTTCAATCCTTT	CGTTTTGGATACGTCTTGGGG	3	29379153 - 29379406
FAN-SCF367	15	20.2	CAGGAGGAAGCTCATGTGGG	TGTGCATGGAACCTAAAGCTCT	3	29119775 - 29119894
FAN-SCF423	16	0.0	ATGGGTGCTGCGTGCTAGCA	CGTTCTCCACGGATCCTAATCC	7	257891 - 257922
FAN-SCF197	16	9.4	CATCAAAAGAGGGGTCCGCT	ACAGTAACCCATGCGATACAGT	1	21455604 - 21455985
FAN-SCF516	16	11.3	TAATCAGACCGCTCCCTCCT	TATGGAGCCCATAGCCAGGT	4	28252096 - 28252225
FAN-SCF286	16	16.9	TGGACGTATGCTGGAGCCTA	GGAGTACTCAGTGCAGCCTG	4	24247781 - 24248036
FAN-SCF553	16	19.9	TGGTCATTTCACTTGTGAATTGT	ACCTTCAAAACAGAAGTTGACA	4	23988922 - 23989203
FAN-SCF98	17	0.0	CAGCAGGAGGTTACTCTGGC	AGCAAGTTAGTAACGCACCT	1	18718021 - 18718404
FAN-SCF917	17	8.9	ACTTGGACCTACCCCTACCC	GACAGGTTAGTGGAATGGCCA	1	17493100 - 17493499
FAN-CON1054	17	13.7	GGTCATCGCTCACCACTTCA	CGCCATAAAATAGTAAGCGGGG	1	15672766 - 15673145
FAN-SCF382	17	18.6	TGCACATAATTGTTGTTTGGCA	GGGTGTGTGTGGTACCATGT	1	12566836 - 12567235
FAN-SCF229	18	0.0	GCACAGGTTTTGGTGCATGT	ACACGACACGACAACACGTA	2	27529617 - 27529951

FAN-SCF194	18	10.6	CTATGGCTGTCTCCGGCAAT	TATGGCGCTTTCGAGACCAA	2	26588193 - 26588592
FAN-SCF354	18	17.0	TCCGTGCGCACATCTATATGA	TAGATGGATTGGGCGCCATG	2	25907743 - 25908140
FAN-SCF872	19	0.0	TCCTGGGATTGTTAGAGTCTGT	CAAAATGCACGTCAGATTATATGA	2	22312544 - 22312930
FAN-SCF89	19	3.9	TGCCCATACTAAAAGTGATCATGG	TGTTCCTGGAGCTTTCATCCA	2	21320143 - 21320464
FAN-SCF5	19	7.1	ACCCAATTCAAGGACCTGCA	TCACCATTACGCATTTACACGA	2	21507575 - 21507885
FAN-SCF178	19	8.8	ACTCGACAATCTGTGGTGGT	TCATGCCCTTAATCGCCCTT	2	21361719 - 21362108
FAN-SCF216	19	9.9	AGCTTCTGAGTACGAAATGCA	ACAACGGATCTTAAAAGAAGGCA	2	21093928 - 21094332
FAN-SCF804	19	10.5	AGCAACCCGAGAAACCTTGT	TGGAGAAGCCTTTTTAATCAAGTG	2	21257000 - 21257326
FAN-SCF359	19	11.0	GTACAGGGGTAAACGCGCTA	CTGGGAGACTGGGATGTTTGA	2	20767369 - 20767760
FAN-SCF385	19	12.8	ACTGTCTTCGTTTCACAACACA	ATGACCTTTCGCTTGCAAGG	2	21702155 - 21702552
FAN-SCF782	19	15.1	AAGAAAAGACCCACCCCACA	ATGGAATCAAAACACTGTGTTCGT	2	21709041 - 21709329
FAN-CON1144	19	16.9	AGCCATGAGTTCTCAATCACTGT	CACGTAAAACCCCGAAACCC	2	21747548 - 21747947
FAN-CON984	20	0.0	AGGAATCATACATTAGTTTCTCCT	TGCTTAGGGAACTGCAAATCCT	3	5608121 - 5608363
FAN-SCF55	20	0.6	AGGGTTGGCCTTGAAATCGA	GACCAGTGCGAACCTCTTGA	3	5867946 - 5868345
FAN-SCF11	20	16.1	GCTTCCAGAACCCTGAAGCT	AGTCAGGCTGTTGGGTAAGG	7	14002188 - 14002310
FAN-SCF625	21	0.0	ACGAAATACTGGCTCTGCTAGG	ACCCCAACAATTCCAAGCCA	7	8259563 - 8259843
FAN-CON1032	21	4.2	TCGTTAAGTGTATGATTCTCTCCA	GTGTGTTAAATTTATGTCGTACCA	7	6515448 - 6515725
FAN-SCF14	21	8.4	CCTCCCTAGCCTCCATCTCA	CTCGATCATCACCTTGGTCCA	2	11289091 - 11289202
FAN-SCF871	22	0.0	TGTGTGACTTTCTGTATGTTCA	CCAGGAACTTGTTGAGGAAAGC	3	9173551 - 9173871
FAN-SCF767	22	3.0	ACATGTTTCTGTGTCAGTTGGT	CCCTTCCGCTGATCTCTGTC	3	7351198 - 7351590
FAN-SCF97	22	6.5	TCCTTTGACATGATTGCCACG	AGTCAATGTCAACCTCCACTGG	3	6958967 - 6959366
FAN-CON1118	22	8.3	GCATTTGCTTTCAAGAGCGC	GCGTAAGTTTGTCCTGCAGG	3	7052910 - 7053309
FAN-CON1141	23	0.0	ACGATTCGACGCCTTTCGAT	ATTGGAGGAGGAGCATGCTT	3	15482868 - 15483267
FAN-CON1063	23	2.2	TCAGAAGAAATATAGCGTGGGT	AAGAATCCCACTCCAACAGCT	3	22688348 - 22688574
FAN-SCF587	23	3.5	GTCTGCAAAAGCAGGCAAGT	AGTCGTTGGATTTTTCTTTTAGGA	3	31471188 - 31471587
FAN-SCF915	23	3.5	CGCGCCAATTGATGAAGGAG	TCGAGCCTGTCATGGTCATG	3	31438822 - 31439221
FAN-SCF793	23	5.7	CCCACCCGAGCCCTTAAAAT	CGTCTGTGCCTATGTCGAGT	3	23487528 - 23487821
FAN-CON1085	23	6.8	GGACCTTGCCTTCCATGGAA	GGTTCTGAAACAAGATCCCCG	3	15926347 - 15926746
FAN-SCF287	23	7.4	TGCATGTGCCGTTTTCACAG	ACCAATGAAACCACGAACCT	3	16140230 - 16140639
FAN-SCF924	23	8.0	TCTGTCGGACAGTCTAAATGG	TGAACCCCTTCTACTGCAAAACT	3	16134694 - 16135091
FAN-SCF78	24	0.0	GACCATTCGTCGCGGATAGT	ACAGCACGTTCTGAGTTGAT	7	23293212 - 23293479
FAN-CON1025	24	6.8	TGAACCGGAAACTCTTACGT	TGGACCAAATCAAAACACACA	7	21504945 - 21505040
FAN-CON1154	25	0.0	GCTGAGTCCCTCGCATGTTA	ACATCACTTCAGCTGCACCA	5	16151073 - 16151358
FAN-CON1130	25	1.2	GCAGGTGGATGTCCGATTCA	GATTTGACGATCGGCCAAGC	5	16155482 - 16155766
FAN-SCF593	25	2.4	CCACTGCTGGAAGCCGAAAC	AGGATGGAATTTATCGCCCGA	4	7038979 - 7039083
FAN-SCF252	25	6.1	ACATGTCAAACGGTGAAAGCA	TATGCGAGCGAGCTAGATGC	3	24661969 - 24662295
FAN-SCF10	26	0.0	GGGCATTGTGGGTGCAAATA	CGGTGATCAAATTTGGGCCA	2	10258308 - 10258711
FAN-SCF458	26	4.2	ACCTAGTTTGAGCTATCTGGCA	CGTCCCACAAGTAGCTAGCT	2	3725132 - 3725214
FAN-SCF126	27	0.0	TCAACAGTTCAACACACTGA	TGTAAGCACTAGAACACTGGA	4	1265833 - 1266095
FAN-SCF164	27	0.0	TGCAGGCATTGGATCGTCAT	TCATTTGCAATGAAGCATCGCT	4	1524501 - 1524913
FAN-SCF390	27	2.4	TCTAGGGTTTGAGGACGCAA	GCATACAATTGGCACGTCCG	4	1770856 - 1771285
FAN-SCF383	27	3.1	CCGTCTCCCATCACTCCTCT	GGCGACTCAAGAGTTACCGA	3	11683723 - 11683764
FAN-SCF520	28	0.0	TGAGGATGTGCAACGTCTCA	TCGGGTTTCCAGATGTCGAC	2	19491481 - 19491897
FAN-SCF896	28	2.5	ATCTGAAGAGGCAAGCAGGG	CCAACTGATCTGCTCCTCCG	2	21512966 - 21513328
FAN-SCF76	29	0.0	GGATGAGAGGAACCGAGGGA	TTGAGATCTCGCACCGTTCC	7	7221386 - 7221749
FAN-SCF743	29	1.2	GCTCCAGCTTAGTCCATGCA	GCCTGAAGAAACACACACCG	4	15153915 - 15154307
FAN-CON1082	30	0.0	TCCGTCCATTGTGAAGAGGC	AGATCAACTTCAGAATCTGAGAA	3	196716 - 196972
FAN-SCF259	30	1.1	TGGGGTTGCACTTGAACAGT	GCGTTTCTTGCTTCAACGGT	3	117480 - 117879

HRM 분석

HRM 반응액은 genomic DNA 10 ng, 10x PCR buffer 2.0 μL, 2.5 mM dNTP mixture 1.0 μL, 0.1 units Taq DNA polymerase (Transgen Biotech, China), SYTO^Ⓡ 9 green fluorescent nucleic acid stain (Life Technologies^TM, USA) 1.0 μL, 각각의 프라이머(Table 1) 10 pmole·μL^-1, 그리고 멸균 된 3차 증류수로 총 20.0 μL로 맞추었다. PCR 반응은 Biometra Tadvanced (Biometra, Germany)를 이용하였으며, 95°C에서 5분간 초기 denaturation을 수행한 뒤, 95°C에서 10초간 denaturation, 60°C에서 20초간 annealing 및 extension 과정을 40회 반복하였다. 마지막으로 72°C에서 15초 동안 full extension 반응을 시켰다. PCR 산물은 LightCycler^Ⓡ Real-Time PCR (Roche, Switzerland)를 이용하여 65°C에서 97°C까지 0.03%씩 온도를 올리면서 각 온도에서 SYTO 9의 형광을 측정하여 melting curve를 그렸고, LightCycler^Ⓡ v.1.1.0.1320 프로그램(Roche, Switzerland)을 사용하여 유전 자형을 분석하였다(Wittwer et al. 2003).

유전자지도 작성

유전자지도 작성은 JoinMap^Ⓡ 4.1 프로그램을 이용하여 수행 하였다(Van Ooijen 2006). 연관 그룹은 LOD≥3.0 조건에서 최대 거리 30 cM을 기준으로 하였고, 지도 거리는 Kosambi (1944) mapping function을 이용하여 계산하였으며, 집단 옵션 은 CP[outbreeder full-sib family, 연관상(linkage phases)을 전 혀 모르고 이형접합(heterozygous)과 동형접합(homozygous)이 불균일하게(heterogeneously) 분포되어 있는 양친을 교배한 집 단]를 사용하였다(Van Ooijen 2011). 최종 유전자 연관 지도는 MAPCHART v.2.1 프로그램을 사용하여 그렸다(Voorrips 2002).

결과 및 고찰

양친의 대량 염기서열 분석

유전자지도 작성용 교배 모본으로 사용된 ‘설향’과 ‘생가생가 나’를 NGS resequencing한 결과, ‘생가생가나’는 총 30.7 Gbp 의 염기서열을, ‘설향’은 총 25.3 Gbp의 염기서열을 해독하였다 (Table 2). 이 염기서열에서 필요없는 부분을 잘라낸(trimming) 결과, ‘생가생가나’는 총 21.8 Gbp, ‘설향’은 총 19.0 Gbp의 염기서열이 남았다(Table 2). 이는 딸기 표준유전체의 길이인 720 Mbp의 약 30배(‘생가생가나’) 및 약 26.5배(‘설향’)에 해당 하는 것이었다(Table 2). 가공된 trimmed reads를 이용하여 딸기 표준유전체인 F. × ananassa (FANhybrid_r1.2) 염기서열에 mapping한 결과, ‘생가생가나’는 총 표준유전체의 약 76%인 530 Mbp를, ‘설향’은 약 76.5%인 534 Mbp를 커버하였다 (Table 2).

Table 2

Statistics of raw reads, trimmed reads, and mapped reads of next generation resequencing data.

Table 2
Plant material	Use	Raw reads	Trimmed reads			Mapped reads

		Total length (bp)	Total length (bp)	Trimmed/ Raw	Genome coverage	The number of mapped reads (%)	Mapped region of reference genome (bp)	Reference genome coverage

‘Senga-Sengana’	Male	30,682,186,222	21,846,702,806	71.2%	≒	193,915,797	530,090,126	76.0%
‘Senga-Sengana’	parent	30,682,186,222	21,846,702,806	71.2%	30.34X	(75.9%)	530,090,126	76.0%

‘Sulhyang’	Female	25,256,529,246	19,080,792,008	75.5%	≒	169,416,290	533,936,123	76.5%
‘Sulhyang’	parent	25,256,529,246	19,080,792,008	75.5%	26.50X	(77.0%)	533,936,123	76.5%

대량 SNP 탐색 및 HRM용 프라이머 디자인

분석된 ‘생가생가나’와 ‘설향’의 염기서열을 비교하여 총 421,480개의 SNP을 탐색하였으며 그 중 양친 모두 동형접합형 인(homozygous) SNP는 206,019개로 가장 많았다(Table 3). 그러나 본 연구에서는 F1 분리집단을 유전자지도 작성용 집단으 로 사용하였기 때문에 사용할 수 없는 SNP였으며 이 SNP를 사용하기 위해서는 F₂ 분리집단을 이용하여야 한다. 그 이유는 F1 집단 모든 개체에서 이형접합형인(heterozygous) 유전자형만 나타나기 때문이다. 나머지 세 가지 경우(동형 대 이형, 이형 대 동형 및 이형 대 이형)의 SNP는 F1 분리집단에서 사용할 수 있다(Table 3). 동형 대 이형 및 이형 대 동형의 SNP는 각각 53,933개 및 74,367개가 탐색되었고, 이 SNP는 F1 분리집단에 서 1:1로 분리되기 때문에 pseudo-testcross mapping에 사용될 수 있는 것이다(Grattapaglia & Sederoff 1994). 이형 대 이형의 SNP는 87,161개가 탐색되었고, 이 SNP는 F1 분리집단에서 1:2:1로 분리되기 때문에 순계를 이용한 F₂ 분리집단과 같은 분리를 보인다. 따라서 본 연구에서는 이 네 가지 경우의 SNP 중에서 이형 대 이형 SNP를 이용하여 유전자지도를 작성하였다 (Fig. 1).

Table 3

Summary of single nucleotide polymorphism (SNP) detection and high resolution melting (HRM) primer design between strawberry cultivars ‘Sulhyang’ and ‘Senga-sengana.’

Table 3
SNP type		The number of polymorphic SNP	The number of non-paralogous SNP	The number of HRM candidates	The number of HRM primer sets designed

‘Senga-sengana’	‘Sulhyang’				Total	Intergenic region	Genic region

							CDSz	Intron	Exon

Homozygous	Homozygous	206,019	8,128	6,750	2,324	1,212	504	416	192
Homozygous	Heterozygous	53,933	2,896	2,347	652	341	134	112	65
Heterozygous	Homozygous	74,367	4,220	3,460	1,338	676	298	247	117
Heterozygous	Heterozygous	87,161	5,604	4,460	1,154	443	458	181	72

Total		421,480	20,848	17,017	5,468	2,672	1,394	956	446

^zCDS indicates a coding sequence region except intron and exon regions.

Fig. 1

A genetic linkage map consisting of 205 single nucleotide polymorphism (SNP) markers using an F₁ sgregating population of strawberry cultivars 'suhyang' x ;Senga-sengana'.

탐색된 총 87,161개의 이형 대 이형 SNP 중에서 non-paralogous SNP의 수는 5,604개였다(Table 3). 본 연구에서 사용한 딸기는 8배체이기 때문에 paralog들이 다수 존재해서 분자표지 개발하 는데 원하지 않는 paralog 부분이 PCR될 수 있기 때문에 이러한 경우를 없애기 위해 non-paralogous SNP만을 선발하였다. 그 중 HRM용 프라이머를 디자인할 수 있는 SNP는 총 4,460개였 고, 실제로 총 1,154개의 HRM 분석용 프라이머를 디자인하여 실험에 사용하였다(Table 3).

HRM 분석용 프라이머를 디자인한 1,154개의 SNP를 유전체 의 위치에 따라 분류해 보면, 유전자 사이(intergenic)에 존재하 는 것이 443개였고, 유전자 내부(genic)에 존재하는 것이 711개 였다(Table 3). 유전자 내부에서도 인트론(intron) 상에 존재하 는 것이 181개, 엑손(exon) 상에 존재하는 것이 72였고, 단백질 로 번역되지 않는 지역(untranslated region, UTR)에 존재하는 것이 458개였다(Table 3).

SNP 분자표지 개발

총 1,154개의 HRM 분석용 프라이머를 디자인하여 SNP 분자 표지 개발에 사용하였다(Table 1, Wittwer et al. 2003). 양친으 로 사용된 ‘설향’과 ‘생가생가나’ 그리고 94개의 F1 개체를 이용 하여 실제로 다형성이 존재하는지 확인하기 위하여 HRM 분석 을 수행하였다. 그 결과 1,154개의 프라이머 조합 중 다형성 (polymorphic)이 나타난 것은 224개였으며, 나머지 930개는 다형성이 없었거나(monomorphic), 두 개 이상의 단편이 증폭되 어 melting curve가 복잡해 유전자형을 구분할 수 없었다. 고추 (Capsicum spp.)의 경우 HRM 분자표지 개발 성공 확률이 80% 인 것에 비하여 훨씬 낮은 성공률(19.4%)을 보였는데(Lee et al. 2013), 단형성(monomorphic)으로 나타난 것은 생물정보 분석에서 거짓양성(false positive)이 선발되기 때문이고, 복잡한 melting curve가 나타난 것은 딸기 표준유전체의 정보가 아직 불완전해 paralog를 완벽하게 제거하지 못하였기 때문인 것으로 생각된다(Hirakawa et al. 2014).

Ge et al. (2013)은 96개의 EST 염기서열로부터 총 116개의 SNP 분자표지를 개발하여 16개의 중국 딸기 재배 품종을 구분하 는데 이용하였는데, 본 연구에서는 NGS resequencing을 이용하 여 이보다 많은 SNP 분자표지를 개발하였다(Table 1). 최근에 Bassil et al. (2015)과 Sargent et al. (2016)은 90 K Axiom^® SNP array를 이용하여 각각 6,594개 및 8,407개의 SNP 분자표 지를 개발하였다. 하지만 이는 high-throughput whole genome genotyping 분석 방법으로 한 번에 9만개의 SNP를 동시에 분석 하는 것이라 본 연구에서 개발한 한 개씩 따로 분석할 수 있는 즉 형질과 연관되어 있다면 육종에 바로 사용할 수 있는 SNP 분자표지와는 다른 것이라 하겠다.

8배체 딸기 유전자지도 작성

개발된 224개의 HRM 분자표지 중 205개는 딸기 유전자 연관지도 상에 위치하였다(Fig. 1, Table 1). 총 연관거리는 800.8 cM이었으며, 8배체 딸기의 경우 28개의 연관군이 작성되 어야 하지만 본 실험에는 30개의 연관군으로 나타났다(Fig. 1). 이는 유전자지도 작성 시 사용된 분자표지의 수가 적었기 때문인 것으로 판단된다. Bassil et al. (2015)과 Sargent et al. (2016)은 총 연관거리가 각각 2,050 cM 및 1,820 cM인 28개의 연관군으 로 구성된 8배체 딸기 유전자지도를 작성한 바 있다. 따라서 본 연구에서 만들어진 유전자지도는 아직 딸기 유전체 전체를 커버하지 못하고 있기 때문에 새로운 분자표지를 유전자지도에 추가해야 할 필요성이 있다. 딸기 유전체 상의 위치를 알고 있는 기 개발된 SSR 또는 SNP 분자표지를 추가함으로써 유전자지도 연관군에 염색체 번호를 부여할 필요도 있다. 하지만 본 연구에서 개발된 SNP 분자표지는 HRM 분석 방법으로 쉽게 분석할 수 있고, non-paralogous SNP 만을 선발하여 개발한 것으로 단인자 멘델 유전에 따라 단순하게 분리하기 때문에 8배체 딸기 유전체 분석 및 다양한 유전 연구에 있어서 큰 도움이 될 수 있을 것이라 생각된다. 또한 본 연구에서 작성한 딸기 유전자지도는 F1 식물체 의 역병 저항성 분석을 통해 QTL 분석에 이용될 수 있을 것이다.

적 요

딸기(Fragaria × ananassa Duch.)는 국내에서 중요한 과채류 작물 중 하나이다. 그러나 현재 재배되고 있는 딸기는 다배체이며 유전체가 복잡하기 때문에 유전체 및 유전학적 연구는 아직 미미한 실정이다. 본 실험에서는 차세대 염기서열 분석(next generation sequencing, NGS) 방법을 이용하여 대량의 SNP 분자표지를 개발하였으며, 이를 이용하여 8배체 딸기 유전자지 도를 작성하였다. 식물재료로는 ‘설향’(모친)과 ‘생가생가나’(부 친)를 양친으로 사용하였고, 이들의 F1 분리집단 94개체를 유전 자지도 작성에 사용하였다. NGS 분석 결과, ‘설향’과 ‘생가생가 나’에서 각각 19.0 Gbp와 21.8 Gbp의 염기서열을 얻었다. 양친 간의 분석된 총 SNP수는 약 87,000개였으며, 그 중 프라이머를 디자인한 SNP의 수는 1,154개였다. High resolution melting (HRM) 분석을 통하여 다형성을 확인한 결과, 1,154개의 프라이 머 중 224개에서만 다형성을 보였으며, 이 중 205개의 HRM 분자표지는 딸기 유전자지도 상에 위치시켰다. 총 연관거리는 800.8 cM이였으며, 30개의 연관군으로 이루어졌다. 본 연구에 서 개발된 SNP 분자표지 및 이를 기반으로 하여 작성된 유전자지 도는 8배체 딸기의 유전체 및 다양한 유전 연구에 큰 도움을 줄 수 있을 것으로 생각된다.

사 사

본 논문은 농림수산식품기술기획평가원 수출전략기술개발사 업(과제번호: 315047-3)의 지원에 의해 이루어졌습니다.

References

1. Bassil N.V., Davis T.M., Zhang H., Ficklin S., Mittmann M., Webster T., Mahoney L., Wood D., Alperin E.S., Rosyara U.R., Putten H.K., Monfort A., Sargent D.J., Amaya I., Denoyes B., Bianco L., van Dijk T., Pirani A., Iezzoni A., Main D., Peace C., Yang Y., Whitaker V., Verma S., Bellon L., Brew F., Herrera R., van de Weg E.. Development and preliminary evaluation of a 90 K Axiom® SNP array for the allo-octoploid cultivated strawberry Fragaria × ananassa. BMC Genomics 2015. 16: 155.
Article
PubMed
PMC
PDF
2. Cox M.P., Peterson D.A., Biggs P.J.. SolexaQA: At-a-glance quality assessment of Illumina second-generation sequencing data. BMC Bioinformatics 2010. 11: 485.
Article
PubMed
PMC
PDF
3. Darrow G.. The Strawberry – History, Breeding and Physiology. New York, USA: Holt, Rinehart and Winston, 1966.
4. Eikemo H., Stensvand A.. Resistance of strawberry genotypes to leather rot and crown rot caused by Phytophthora cactorum. Eur. J. Plant Pathol 2015. 143: 407-413.
Article
PDF
5. Folta K.M.. Functionalizing the strawberry genome – a review. Int. J. Fruit Sci 2013. 13: 162-174.
Article
6. Folta K.M., Davis T.M.. Strawberry genes and genomics. Crit. Rev. Plant Sci 2006. 25: 399-415.
Article
7. Ge A.J., Han J., Li X.D., Zhao M.Z., Liu H., Dong Q.H., Fang J.G.. Characterization of SNPs in strawberry cultivars in China. Genet. Mol. Res 2013. 12: 639-645.
Article
PubMed
8. Grattapaglia D., Sederoff R.. Genetic linkage maps of Eucalyptus grandis and Eucalyptus urophylla using a pseudo-testcross: mapping strategy and RAPD markers. Genetics 1994. 137: 1121-1137.
Article
PubMed
PMC
PDF
9. Hirakawa H., Shirasawa K., Kosugi S., Tashiro K., Nakayama S., Yamada M., Kohara M., Watanabe A., Kishida Y., Fujishiro T., Tsuruoka H., Minami C., Sasamoto S., Kato M., Nanri K., Komaki A., Yanagi T., Guoxin Q., Maeda F., Ishikawa M., Kuhara S., Sato S., Tabata S., Isobe N.. Dissection of the octoploid strawberry genome by deep sequencing of the genomes of Fragaria species. DNA Res 2014. 21: 169-181.
Article
PubMed
10. Isobe S.N., Hirakawa H., Sato S., Maeda F. Ishikawa M., Mori T., Yamamoto Y., Shirasawa K., Kimura M., Fukami M., Hashizume F., Tsuji T., Wada T., Ono A., Kawashima K., Nakazaki N., Kishida Y., Kohara M., Nakayama S., Yamada M., Fujishiro T., Watanabe A., Tabata S.. Construction of an integrated high density simple sequence repeat linkage map in cultivated strawberry (Fragaria × ananassa) and its applicability. DNA Res 2013. 20: 79-92.
PubMed
11. Jung S., Cestaro A., Troggio M., Main D., Zheng P., Cho I., Folta K.M., Sosinski B., Abbott A., Celton J., Arús P., Shulaev V., Verde I., Morgante M., Rokhsar D., Velasco R., Sargent D.J.. Whole genome comparisons of Fragaria, Prunus and Malus reveal different modes of evolution between Rosaceous subfamilies. BMC Genomics 2012. 13: 129.
Article
PubMed
PMC
12. Kim J.E., Oh S.K., Lee J.H., Lee B.M., Jo S.H.. Genome-wide SNP calling using next generation sequencing data in tomato. Mol. Cells 2014. 37: 36-42.
Article
PubMed
PMC
13. Kosambi D.D.. The estimation of map distance from recombination value. Ann. Eugen 1944. 12: 172-175.
14. Lee J., Park S.J., Do J.W., Han J.H., Choi D., Yoon J.B.. Development of a genetic map of chili pepper using single nucleotide polymorphism markers generated from next generation resequencing of parents. Hortic. Sci. Technol 2013. 31: 473-482.
Article
15. Lerceteau-Köhler E., Guérin G., Laigret F., Denoyes-Rothan B.. Characterization of mixed disomic and polysomic inheritance in the octoploid strawberry (Fragaria × ananassa) using AFLP mapping. Theor. Appl. Genet 2003. 107: 619-628.
Article
PubMed
PDF
16. Li H., Durbin R.. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 2009. 25: 1754-1760.
Article
PubMed
PMC
PDF
17. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, Marth G, Abecasis G, Durbin R. The sequence alignment/map format and SAMtools. Bioinformatics 2009. 25: 2078-2079.
Article
PubMed
PMC
PDF
18. Monfort A., Vilanova S., Davis T.M., Arús P.. A new set of polymorphic simple sequence repeat (SSR) markers from a wild strawberry (Fragaria vesca) are transferable to other diploid Fragaria species and to Fragaria × ananassa. Mol. Ecol. Notes 2006. 6: 197-200.
Article
19. Rousseau-Gueutin M., Lerceteau-Köhler E., Barrot L., Sargent D.J., Monfort A., Simpson D., Arús A., Guérin G., Denoyes-Rothan B.. Comparative genetic mapping between octoploid and diploid Fragaria species reveals a high level of colinearity between their genomes and the essentially disomic behavior of the cultivated octoploid strawberry. Genetics 2008. 179: 2045-2060.
Article
PubMed
PMC
PDF
20. Sargent D.J., Davis T.M., Tobutt K.R., Wilkinson M.J., Battey N.H., Simpson D.W.. A genetic linkage map of microsatellite, gene-specific and morphological markers in diploid Fragaria. Theor. Appl. Genet 2004. 7: 1385-1391.
Article
PDF
21. Sargent D.J., Fernandéz-Fernandéz F., Ruiz-Roja J.J., Sutherland B.G., Passey A., Whitehouse A.B., Simpson D.W.. A genetic linkage map of the cultivated strawberry (Fragaria × ananassa) and its comparison to the diploid Fragaria reference map. Mol. Breed 2009. 24: 293-303.
Article
PDF
22. Sargent D.J., Passey T., Šurbanovski N., Lopez-Girona E., Kuchta P., Davik J., Harrison R., Passey A., Whitehouse A.B., Simpson D.W.. A microsatellite linkage map for the cultivated strawberry (Fragaria × ananassa) suggests extensive regions of homozygosity in the genome that may have resulted from breeding and selection. Theor. Appl. Genet 2012. 124: 1229-1240.
Article
PubMed
PDF
23. Sargent D.J., Yang Y., Surbanovski N., Biance L., Buti M., Velasco R., Giongo L., Davis T.M.. HaploSNP affinities and linkage map positions illuminate subgenome composition in the octoploid, cultivated strawberry (Fragaria × ananassa). Plant Sci 2016. 242: 140-150.
Article
PubMed
24. Shulaev V, Sargent DJ, Crowhurst RN, Mockler TC, Folkerts O, Delcher AL, Jaiswal P, Mochaitis K, Liston A, Mane SP, Burns P, Davis TM, Slovin JP, Bassil N. The genome of woodland strawberry (Fragaria vesca). Nat. Genet 2011. 43: 109-116.
PubMed
25. Van Dijk T., Pagliarani G., Pikunova A., Noordijk Y., Yilmaz-Temel H., Meulenbroek B., Visser R.G., van de Weg E.. Genomic rearrangements and signatures of breeding in the allo-octoploid strawberry as revealed through an allele dose based SSR linkage map. BMC Plant Biol 2014. 14: 55.
PubMed
PMC
26. Van Ooijen J.W.. JoinMap® 4, Software for the calculation of genetic linkage maps in experimental populations. Wageningen, the Netherlands: Kyazma B.V., 2006.
27. Van Ooijen J.W.. Multipoint maximum likelihood mapping in a full-sib family of an outbreeding species. Genet. Res 2011. 93: 343-349.
Article
28. Vilanova S., Sargent D.J., Arús P., Monfort A.. Synteny conservation between two distantly-related Rosaceae genomes: Prunus (the stone fruits) and Fragaria (the strawberry). BMC Plant Biol 2008. 8: 67.
Article
PubMed
PMC
PDF
29. Voorrips R.E.. MAPCHART: Software for the graphical presentation of linkage maps and QTLs. J. Hered 2002. 93: 77-78.
Article
PubMed
30. Wittwer C.T., Reed G.H., Gundry C.N., Vandersteen J.G., Pryor R.J.. High-resolution genotyping by amplicon melting analysis using LCGreen. Clin. Chem 2003. 49: 853-860.
Article
PubMed
PDF

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A genetic linkage map of allo-octoploid strawberry (Fragaria × ananassa Duch.) using SNP markers

Korean. J. Breed. Sci.. 2017;49(3):119-128. Published online September 1, 2017

DOI: https://doi.org/10.9787/KJBS.2017.49.3.119

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A genetic linkage map of allo-octoploid strawberry (Fragaria × ananassa Duch.) using SNP markers

Korean. J. Breed. Sci.. 2017;49(3):119-128. Published online September 1, 2017

DOI: https://doi.org/10.9787/KJBS.2017.49.3.119

Figure

A genetic linkage map of allo-octoploid strawberry (Fragaria × ananassa Duch.) using SNP markers

Fig. 1 A genetic linkage map consisting of 205 single nucleotide polymorphism (SNP) markers using an F1 sgregating population of strawberry cultivars 'suhyang' x ;Senga-sengana'.

Fig. 1

A genetic linkage map of allo-octoploid strawberry (Fragaria × ananassa Duch.) using SNP markers

Table 1

List of HRM primer sets used in this study.

Marker name	LG	Position (cM)	Forward primer	Reverse primer	Fragaria vesca genome

					Chromosome	Position (bp)

FAN-SCF420	1	0.0	GGAATACCGAGACTCGTCACT	ACGACAATCCTATTCAATCCACTG	6	30689884 - 30690283
FAN-SCF813	1	9.0	TGAATAAAGGGTTTTCAGTTTTGA	TCAGACATCTCTCAAGGTCGT	6	30647220 - 30647488
FAN-SCF384	1	10.2	ACCAACAACTCAAACTTCAAAAGT	GGCATTTTGTTTTATGACGAGACT	6	31592825 - 31593037
FAN-SCF132	1	11.4	AGTGGAGGACATATTCAAGTGTCT	GTTGCGGTGCTTAAGAGTGC	6	31401845 - 31402250
FAN-SCF418	1	12.8	TCGAAATTCACACGAAGAAGA	TTGGACTATACTCTGCAATACTG	6	31347505 - 31347862
FAN-SCF892	1	15.4	TGGGCCACTCTGTCTATGTT	TCGGCTTGTCCATCCTTTCC	6	31458344 - 31458618
FAN-CON1139	1	21.4	TGCAGGGGACTCTGAGATCA	TGGAAGTCCAGGAGTACTTGG	6	28464183 - 28464582
FAN-SCF833	1	38.6	TGAAGTGCTGCAATCCTGGT	AACGGTGGGCTAGATTTGGG	6	25589098 - 25589497
FAN-SCF174	1	66.7	TGTTGTAGTGAGAGCCGTCG	ACCAAAGGGCGATGATCCTC	6	15978363 - 15978658
FAN-SCF715	1	71.4	TGAAGGAAGAGGAGGAAGTGT	TGGTTTAGGGTGTTTTAGGGGT	6	14511815 - 14512028
FAN-CON1046	1	78.9	CTAGCCTGGTCAACCGACTC	TCCCATTAGTGTGGTTTAGCGA	6	10237152 - 10237439
FAN-SCF338	1	102.0	TGCATACATTTTCTGTGCGA	CCAGGTAACGATAAGGGCCC	6	7114971 - 7115361
FAN-SCF735	1	103.8	TGCGACAGGATAATCATGACGT	ACCCCTACTCCATCATCCCC	6	7158469 - 7158868
FAN-SCF858	1	113.4	AGAATCTAGACTGATGGATCGT	ACCGACGTAATAACAACCACCA	6	4581492 - 4581890
FAN-SCF33	2	0.0	CTACTGCCCACAACTCGGTG	CCAATGGCTACAATCTGCGC	7	16926268 - 16926609
FAN-SCF607	2	68.5	CTCGTGAAATTAGCATCCGTCA	TCTCTATGCACACTGACAACCA	4	24082358 - 24082757
FAN-SCF377	2	70.8	GCTGATGAAACATAGTAATTCCGA	TGCGGACAAAATGGATATTTTCT	4	26910570 - 26910801
FAN-SCF468	2	72.0	ACCAAAATCATAAGCAAAACCAGA	TCAGAAGGAAGCTTTCTAGAGCA	4	26624055 - 26624454
FAN-SCF399	2	72.0	ACCTCTCAAATGTGGTTCTCA	CCCTCTAGGACCTTCTGATGC	4	26707960 - 26708303
FAN-SCF405	2	72.0	TCCACAAGAACCTAAAAGCAT	GGTGAAGGTAAATGAAAAATCTGT	4	26145596 - 26145936
FAN-SCF719	2	72.0	AGCCTCCCAGCCTAACAGTA	CATGATTCAATGACGCGGCC	4	26613015 - 26613412
FAN-SCF39	2	73.1	CATGGCAGGCTAGCTTCGAT	TGGACCGACTTCCTCAACAG	4	26359016 - 26359415
FAN-SCF52	2	74.7	TGCTCTTTCGAAATGCCAGTG	TCACACACTGCTTAGCAACTGA	4	26652718 - 26653117
FAN-SCF956	2	77.0	ACTCTTAGCTAAATGGTTCTTGCT	ACCCTATATTGCAAGAACACCT	4	24693600 - 24693962
FAN-SCF824	2	79.2	TTCCCTATATATGTTTCCCATGCA	ATGCGCACCACCGAGCTTAG	4	24568435 - 24568834
FAN-SCF309	2	80.8	CCAATAAAACTTGCGGCGGT	TGGTGGGTGCAATTTGATTG	4	24851133 - 24851532
FAN-SCF637	2	81.9	ACTAGGAGGAAGGGGCCTTT	ACACCAAGATCAGCATCATCA	4	26476579 - 26476858
FAN-SCF578	2	86.0	GCTATTCAGCGGGGTAAGCA	GCTTAGTGTCAAAAATGCAGCC	4	25972133 - 25972532
FAN-CON1146	2	88.7	AGTCTCCAAAGCCTGAAACCA	CCAGGTATTCGGTCAGTTGCT	4	25186628 - 25187027
FAN-SCF6	2	89.2	TCGCAAATGGCTTTTTAGTTGC	AGACTGTATTTGTCTGAGACGA	4	25176196 - 25176558
FAN-SCF131	2	89.8	ACAATCAAACGTGAGGGGCT	CCAACGATGAATGTGAAGAACAGA	4	25687752 - 25688153
FAN-SCF909	2	94.5	ACCCCTAACTAAGTCATCACCG	CCTTGTTGGGTCTGGGCTAA	6	21985116 - 21985169
FAN-SCF945	2	96.7	TGCTGCACATTGTTCAGTCT	TGTTTATTCAGTGTTCATCAGATG	4	23040342 - 23040504
FAN-SCF595	2	98.5	TGGTGCAGGATGAGAATGAGG	CACCTGGTGACGGTCATTCA	4	22850917 - 22851309
FAN-SCF345	2	102.6	AAGGGGACGGTACAGCTACT	GGTGCCATCCATTGAGGCTA	4	22292764 - 22293102
FAN-SCF163	2	103.9	GGTTCGCTATGGCTAGGTCC	AGCTCAAGTTGTGGGTTCCA	4	22363717 - 22364056
FAN-SCF231	2	104.5	ACTGCTTTCCTCTGTGGTTT	AGAGGCATACAACACTGAGTCT	4	22008076 - 22008475
FAN-SCF294	2	106.8	GTGCCTTCAAGGAGCTGGAT	CCCTGCCATAAATTGTTAAATGCA	4	22202764 - 22203163
FAN-SCF402	2	109.1	GCACGCCTCCTCAGTTACAT	CGAGCTGACGTTTAAGTCCCA	4	22640567 - 22640947
FAN-SCF108	3	0.0	AGGGTGTGGAATATCATCACTCA	ATGGGCCTTTGTACCGGTTT	6	21857853 - 21858252
FAN-SCF389	3	22.4	TCTAGCGCAATCTGCACCAA	TCAATGAAACGTACGGCGGT	6	21995951 - 21996362
FAN-SCF166	3	26.1	TACACCGACAGGGAGGTTGA	TTGATCCGAGCGAGCTTACC	6	22152785 - 22153184
FAN-SCF59	3	26.1	GGCTCTGACTTCGGCTTCTT	TCTGCTGGCATTTTGTGTGC	6	22245209 - 22245570
FAN-SCF51	3	26.1	CAGGAGGGACTGGAAAGACA	TGTGTTGTCATCCCACTAAGCA	6	22330843 - 22331242
FAN-SCF135	3	26.7	TCGTTGGAACCTCAGGTTGG	AGGGATATTGATGATATTGTTGCA	6	21563283 - 21563682
FAN-SCF185	3	27.2	GTGCTACCGCTGAGGACAAT	CTAACATCCTCCTGGGCGTG	6	21786636 - 21787035
FAN-SCF105	3	27.2	TTTCGCTTCCAAGTCACCAAC	AGTTTGTCAAAATCTCACCTCC	6	21410739 - 21411138
FAN-SCF215	3	27.8	CGTCGTAGTGGATGATCCTT	TGACAAATGACAACCCATGTT	6	22851692 - 22852092
FAN-SCF58	3	28.3	TGGGGTAAGTGCATTTGGTCA	CCAAAGCCACTCCTCTTCGT	6	23743188 - 23743587
FAN-SCF83	3	28.3	ATGGGCTCCAGCTCTAGTCA	AGAGAGAGTTCAAAAGAGGGGT	6	23220621 - 23221030
FAN-SCF115	3	28.3	GGTGGACATGAATATGAGAAAGCA	GGAAAGACCCCTACAAAAACCC	6	23209827 - 23210225
FAN-SCF173	3	28.9	ACCAGACTACCACTGTGCCA	AAGCAGGGTCGACAACTACC	6	23136810 - 23137047
FAN-SCF388	3	30.7	ACCCTGCCTCCTGTTCAAAC	CACGGCTATACCCAAAGAGCT	6	24286158 - 24286551
FAN-SCF437	3	33.2	TTCTTGCAGGCAGACCAGTC	GGTGCACTGAGGTGGTTTCT	6	22909018 - 22909416
FAN-SCF238	3	33.8	AGCTGTCTAAAAAGCTCTTCGGA	TCGCAACAGAAGTGACTCCC	6	21833742 - 21834148
FAN-SCF450	3	33.8	CCGAAACATGGTCGATCCGT	CCCAACCTTGCTGCTATTGC	6	22164873 - 22165272
FAN-CON1153	3	34.4	CCCATACGCGTGGCTTCAAT	ACTTATCCTCTTCTTTGATGTGGG	6	21433559 - 21433813
FAN-SCF438	3	43.0	TTCCCAGATCACTCTGCTGA	GGGTTGTTAAGGTATAGTACTTGG	6	13401292 - 13401547
FAN-SCF436	3	47.2	ACCAATTTATCGAAACTTCACCGA	TCGATCAAAGACTTGCTATCGA	6	14629000 - 14629399
FAN-SCF246	3	48.3	TGAGGAGTAGGAGCTGTCCA	TTAGCTAAGCAGGGTTGGCG	6	10601014 - 10601413
FAN-SCF165	3	50.1	GCACAAAATATTGCTGATTAGGCA	TGCAGTTTCTTATATGTTGTACGG	6	14765076 - 14765398
FAN-SCF79	3	59.0	TGTTGTTTCTTCCTCCTTGGT	AGTTGGTAAAAGTCTAGGTATGGA	6	11024227 - 11024401
FAN-SCF233	3	60.1	CCAACTCAAATGTGAAGTTGTGGA	TCCGCCCAGACTCAAAACTT	6	9124886 - 9125097
FAN-CON1070	3	65.5	GCAAGCTTTGGCACTCATCG	TCCCTCAGGAGATGGTTGGT	6	9045076 - 9045396
FAN-SCF443	4	0.0	TCACAACGCTTGTCAAAGGC	ACCAGAAAGAGTATCCTCGTCT	-	-
FAN-SCF172	4	3.5	TTGAGGCTCATTGACAACGT	TGAGGGAGAGAGATAGAGGTGC	6	3493659 - 3493907
FAN-SCF316	4	6.5	ACAGCCTCAATCAGCTGCTT	TAAGTTTGGGCGATGATCCT	6	2504998 - 2505390
FAN-SCF769	4	30.7	CCGAAGAGATGAGCAACGGT	GGGACTTGATCATTTGCCGG	6	9394367 - 9394762
FAN-SCF656	4	32.5	ACATATCGCTGCTGGACCAC	AGCTAGCAACTTCCGACCAA	6	10822978 - 10823285
FAN-SCF119	4	34.6	CCATCATTTTACAAACGCTACACA	ACGTGATCTTGCAACGGGAA	6	11132414 - 11132696
FAN-SCF569	4	35.3	TGACTTTTCGTCCATGTTGTCA	GACCGAAACGTATCCATAAATAAA	6	10946444 - 10946849
FAN-SCF570	4	37.7	ACGAAAGTGGAGGTGCTCTG	TGAGCGAGATGAAAACGTCG	3	5315532 - 5315808

FAN-SCF273	4	43.8	TCTCAAACGTCTTGACGCCT	TGAGTGCTATTGTGGCGACT	6	18744174 - 18744235
FAN-CON1062	4	43.8	ACACCACGATATGCAACCACT	ATTTCAAATGGGCCGGGTCA	6	14238062 - 14238401
FAN-SCF183	4	44.3	ACCATCGACGATGAGGAAGC	CGGGGATGCTGGTCATACTT	3	31695069 - 31695130
FAN-SCF69	4	48.8	AGCGGTCCATTGACTCAACA	TGTTGTAAAGCTTGTCGGGA	6	14700987 - 14701320
FAN-CON1071	5	0.0	TTGAGGTTTAAGCCGAGATGACT	GTACTAACAGAGGCGGCGTT	6	27458588 - 27458625
FAN-CON1087	5	6.2	TGCAGATGTTGACTCTGCGT	TTTTTACGTCCGCCGACTGA	4	32415681 - 32415732
FAN-SCF290	5	7.3	TTCACTCAAAAATTCGTGTGTT	ACTTCTTACACTGACAAAACTCGA	4	32269269 - 32269656
FAN-SCF168	5	35.6	TCAACAGAGAAGATGCTTGGT	ACACTCCCTTTTCCTAGCACA	6	9735998 - 9736029
FAN-CON1123	5	37.6	TCGTCACATCTACAATTGGCTCT	GCGATGGTTCTAAGGTGTTCG	4	23313914 - 23314024
FAN-SCF965	5	39.3	TGTGGAGAAGTTCTAAGAATCCCG	CGATCAAAGATGAAACCTAATTCC	6	1903611 - 1903648
FAN-CON1056	5	42.8	AGTGATTGGGCTGCATCAAA	GCTAGGTACAGTTGCCCTGG	4	21935175 - 21935548
FAN-CON1143	5	47.0	TGGTAGCCTTCCCAATGCAG	ATAAGCCAACCCCCGAGTTG	5	11569758 - 11570157
FAN-CON1128	6	0.0	TCGACATAGTGAACGGTGGC	CAAATGGATTTGCGGCACCA	7	423374 - 423768
FAN-CON1053	6	1.1	GCTGCAACAAAATTGGCTTCC	AGGAGACTCTAAGGTATTGATCGA	7	344631 - 344903
FAN-CON1059	6	2.3	CCTAGCTCGGCAATCAGAGG	TGCAGTCAGGAGCAGTTGAG	6	30104705 - 30104918
FAN-CON1097	6	15.3	ACGTTGACGCGTAATTTGGA	TGCAGTTGGTGAATGTTTGA	7	6499804 - 6500187
FAN-SCF474	6	19.3	TCGTGGGCTTAAGTTCTCAAGA	CTCGCCAAGAGTATTCTCCA	-	-
FAN-CON1122	6	21.0	AGGACTTCTTTGATAGTACCGACA	TCTTCCCAACCCACAAGCAA	7	7598690 - 7599115
FAN-SCF145	6	23.4	GCAATTTCTAGAGAGGCAAGCA	TGCTTTTAAAGACAGAGAAGACA	7	5750781 - 5750840
FAN-CON1073	6	33.3	ACCCGTGAAACAATAGGGAAC	AAAGCGATATACATAACATGCACC	7	11248187 - 11248367
FAN-SCF961	6	34.6	TGTGCGAATTTAATTTCCACACA	GAGGAAGCATGAGCATCGGA	1	2921841 - 2921960
FAN-SCF139	6	41.1	TGCAACTCAACCAAGCCAGA	AGTTCACACTTTGTAGCCACTGA	7	12594627 - 12594928
FAN-SCF730	7	0.0	ACCGGGTAACTTTGTTGGCA	ACTCACCAATATCCGATGTGGT	2	10795356 - 10795437
FAN-CON1134	7	1.7	ATCTGAGGTTTTAATTCTCATGC	TGGTATTGTACAAGGAAAGAGAGG	7	2111414 - 2111539
FAN-CON1145	7	2.8	TGGAAGCACTAGCCTTTGTCA	GCAAAAGGAAAGAAATAACGTGGG	5	8070408 - 8070518
FAN-CON1115	7	23.5	TCATTTCCTGGCAGCTACTGA	TCAGAAACCAAACTAGCCAAT	5	11182526 - 11182760
FAN-SCF756	7	32.7	GCTACTGCATTTTAGCACTTTGC	TTGATAAACCAGTCTTATAACGCG	5	24691171 - 24691218
FAN-CON1066	8	0.0	CCTTTTGGGCCCATGACTTG	GGTGTGATTAATCGGCCCGA	5	8945836 - 8946223
FAN-SCF378	8	0.0	CAAGAGGCAGCTTCTCTGGT	GACTCCACAAAGCCTGGTCA	5	9274286 - 9274540
FAN-CON1069	8	9.7	TTGTCGTCTTAATCAAATGATTGT	ACAGCTATATCCTCCACATGT	5	5735911 - 5736310
FAN-SCF698	8	12.0	GCCGAAAACCGTGGTTTGAG	ACCGTGCTTGTTGTACTCGT	6	4563039 - 4563335
FAN-SCF27	8	12.5	CTGTGACTCTGTGAGGGCAG	TGCAGACATGAAATGCACCA	5	5272835 - 5273234
FAN-CON1117	8	15.4	GTTTCACCATCCAAGCTGCG	TCAACCCACAACCACCAACT	5	4918889 - 4919285
FAN-SCF227	8	16.0	TCACCGAGAGTAGCCTAGGA	AGATGCCAGGTTACTAGTTACT	5	4858458 - 4858855
FAN-SCF63	8	25.0	GTTCTGATTGCTAGCATCTGCA	TCCTGATGCGCTACTCTTGT	4	32169627 - 32169969
FAN-SCF480	8	27.3	AGGATAACAATCTGTAGTGGGGA	TCACCTTAGGCCTCAGCAAA	5	2964486 - 2964724
FAN-CON1015	8	30.2	AGATTGTTGGACGCCTCTAGTC	TCCTCATCAGATAAGGTGTGCA	5	1708322 - 1708548
FAN-SCF260	8	30.8	TCTCTTGAACACATTGAACCACA	TCTATAATCGGGTTCTTGGGT	5	1188138 - 1188537
FAN-SCF434	9	0.0	ACGTTGGTTTCTAAGGGTGT	CGATCACAAGGTCCTGAGAGA	6	35969226 - 35969306
FAN-SCF279	9	2.2	CCACCTGGGGGTTGAAAGTA	CCGCTTTGCATAACTTTCGGT	2	15447700 - 15447757
FAN-SCF326	9	2.9	TGGTTTTTCGCATTTGTTGGT	GGGGACGCTTGAAGAACACA	6	2118937 - 2119014
FAN-SCF846	9	7.7	TGTCAGACACAAGTTAGCTTTTGA	AATCACGGTCCAACACTCGT	6	35292089 - 35292279
FAN-SCF847	9	18.4	GGAGCTTCTTTATTTTTACTCGCA	TGCAAAATCCGGACAGCAAC	6	32014064 - 32014193
FAN-SCF88	9	26.8	TCATACTTGTTGCTGTAGTCACA	GGGTTCTGTTTGTGGTTGGG	6	17704328 - 17704727
FAN-CON1099	9	30.6	AGGAGAAGGAATTGGATCACA	AGGTTCTTTCTTGGACTGGCA	6	28740651 - 28740822
FAN-SCF369	10	0.0	TCAAGTTCACGTTTTACAGTGTCA	TGCATTCTCACCTTCTCAACT	2	14150750 - 14151150
FAN-CON1089	10	15.0	GGCACAACCTGTTTGAGTTT	AGTCCATTCTGCTCCAGTCT	2	3613082 - 3613418
FAN-CON1105	10	19.8	GCGATGACCCAATTTTCGGG	GCCACATCTGTCACATATGCC	2	10339151 - 10339546
FAN-SCF154	10	20.4	TGCCAGCGAATGAGTAGCAG	ACTGCTTATTTGCTTTGGCCA	2	10167626 - 10168044
FAN-SCF559	10	21.0	GCAAATTGAGTACAGTGACCGA	CCTTGGGTTTGCCTGTAGAA	2	10088172 - 10088571
FAN-SCF1	10	29.6	GATACAGACAGGTTTGTGCGC	TATGGTCTCGCAGAAGCCAC	2	10333513 - 10333912
FAN-SCF605	11	0.0	TGCCCCTCCTCTCGTTTTTC	TGACTTCGAACTTGACCTCCA	6	18216050 - 18216445
FAN-SCF807	11	18.9	TGCAATAAAGCAAAGTTCAAAAAT	AGTTCTTTTGCTTCGTACACT	6	28062711 - 28062898
FAN-CON1016	11	24.9	AGGGAATGGAGGTATAAGAGTT	CCTTTCTGGTAGCAGCCTTACT	6	26052420 - 26052541
FAN-SCF67	12	0.0	AAGGGTGGTTTTAGGGCTGG	CGGGCTTTCACCGATCTCTA	4	7444429 - 7444493
FAN-SCF203	12	1.6	ATTTGCGAACCTGTGACCAC	AGAAAATGATGGCGCTTCAACA	3	8554786 - 8554880
FAN-SCF235	12	6.6	ACCCACTATGGAGTTAGCTCT	TCCAACAAACACCTAGGGCA	6	3571899 - 3572282
FAN-SCF129	12	11.4	TGGCGATACAAGTTCGTAAACA	AGTGAGGCTGATTTAGAGTTTCCA	5	14764214 - 14764422
FAN-SCF2	12	23.9	TCGAGAGGTGTATAAAATGCTCT	TGTTTGTATGCATAACAAGCTGA	4	5849067 - 5849236
FAN-SCF548	13	0.0	CCTGGGGTGTTCAACTAGGC	ATCTGCCGTGTGATGCAAGA	5	10208591 - 10208636
FAN-SCF112	13	5.1	TGTAGGAGACAAAGCGTAGTGA	TGAGTAGCAGCAGACTGTAACA	6	11944125 - 11944534
FAN-SCF34	13	5.7	ATTTGAAACGCGTGGATCGAG	GTCTCCAGCACCACCAGGAA	2	1358158 - 1358239
FAN-SCF84	13	7.3	CTTGAGGCCGACGATCTGTT	AGATTCTGCCATCCCGCATC	3	1653607 - 1653678
FAN-SCF483	13	12.8	GGGCATTGGGATCTCAGCTT	CGTCGCCTAGTGACAAGGAA	6	9565771 - 9566099
FAN-SCF760	13	15.1	CAATGACCCTCACTCACCCC	ACCCTACACCATGATCCTCCA	6	9911049 - 9911186
FAN-SCF263	13	22.8	TGTTTAACGGATCATAAATGCAT	GTCATTCGCGACTCGGTACA	6	12241307 - 12241731
FAN-CON1039	14	0.0	GAGATGAGGATGCAGGCCAA	TGCTCACTCTATCGATGGGA	6	20401011 - 20401336
FAN-CON1142	14	6.8	AGTTCATCCGAAGCTGTGCA	ACTGGATATACCGCGCATGT	6	12579743 - 12579956
FAN-CON1137	14	10.9	TCACCTGGAAATTCAGAGGA	TGCCAAAGGTCGAAAATGCG	6	13012521 - 13012836
FAN-SCF289	14	22.2	ACCCTGAAATGGTTAACGTGGT	GCCATATGCCTTACTCATATCT	6	20589193 - 20589212
FAN-SCF905	15	0.0	GGAGGGCTGTTCTTCAGTGG	TCTTACGCAGTGCAGTCCTG	3	22549713 - 22549953
FAN-CON1076	15	1.2	GGTGCAATCCAATGAGCAGG	ACTCAGGCTCGGTAGACCAT	3	22549227 - 22549626
FAN-SCF362	15	6.1	CTGGATCGCTACCTGCACTC	TTAGGAGGGGTCTTCCGTGA	2	6880598 - 6880965
FAN-SCF66	15	17.8	TCCCAGCGCTTCAATCCTTT	CGTTTTGGATACGTCTTGGGG	3	29379153 - 29379406
FAN-SCF367	15	20.2	CAGGAGGAAGCTCATGTGGG	TGTGCATGGAACCTAAAGCTCT	3	29119775 - 29119894
FAN-SCF423	16	0.0	ATGGGTGCTGCGTGCTAGCA	CGTTCTCCACGGATCCTAATCC	7	257891 - 257922
FAN-SCF197	16	9.4	CATCAAAAGAGGGGTCCGCT	ACAGTAACCCATGCGATACAGT	1	21455604 - 21455985
FAN-SCF516	16	11.3	TAATCAGACCGCTCCCTCCT	TATGGAGCCCATAGCCAGGT	4	28252096 - 28252225
FAN-SCF286	16	16.9	TGGACGTATGCTGGAGCCTA	GGAGTACTCAGTGCAGCCTG	4	24247781 - 24248036
FAN-SCF553	16	19.9	TGGTCATTTCACTTGTGAATTGT	ACCTTCAAAACAGAAGTTGACA	4	23988922 - 23989203
FAN-SCF98	17	0.0	CAGCAGGAGGTTACTCTGGC	AGCAAGTTAGTAACGCACCT	1	18718021 - 18718404
FAN-SCF917	17	8.9	ACTTGGACCTACCCCTACCC	GACAGGTTAGTGGAATGGCCA	1	17493100 - 17493499
FAN-CON1054	17	13.7	GGTCATCGCTCACCACTTCA	CGCCATAAAATAGTAAGCGGGG	1	15672766 - 15673145
FAN-SCF382	17	18.6	TGCACATAATTGTTGTTTGGCA	GGGTGTGTGTGGTACCATGT	1	12566836 - 12567235
FAN-SCF229	18	0.0	GCACAGGTTTTGGTGCATGT	ACACGACACGACAACACGTA	2	27529617 - 27529951

FAN-SCF194	18	10.6	CTATGGCTGTCTCCGGCAAT	TATGGCGCTTTCGAGACCAA	2	26588193 - 26588592
FAN-SCF354	18	17.0	TCCGTGCGCACATCTATATGA	TAGATGGATTGGGCGCCATG	2	25907743 - 25908140
FAN-SCF872	19	0.0	TCCTGGGATTGTTAGAGTCTGT	CAAAATGCACGTCAGATTATATGA	2	22312544 - 22312930
FAN-SCF89	19	3.9	TGCCCATACTAAAAGTGATCATGG	TGTTCCTGGAGCTTTCATCCA	2	21320143 - 21320464
FAN-SCF5	19	7.1	ACCCAATTCAAGGACCTGCA	TCACCATTACGCATTTACACGA	2	21507575 - 21507885
FAN-SCF178	19	8.8	ACTCGACAATCTGTGGTGGT	TCATGCCCTTAATCGCCCTT	2	21361719 - 21362108
FAN-SCF216	19	9.9	AGCTTCTGAGTACGAAATGCA	ACAACGGATCTTAAAAGAAGGCA	2	21093928 - 21094332
FAN-SCF804	19	10.5	AGCAACCCGAGAAACCTTGT	TGGAGAAGCCTTTTTAATCAAGTG	2	21257000 - 21257326
FAN-SCF359	19	11.0	GTACAGGGGTAAACGCGCTA	CTGGGAGACTGGGATGTTTGA	2	20767369 - 20767760
FAN-SCF385	19	12.8	ACTGTCTTCGTTTCACAACACA	ATGACCTTTCGCTTGCAAGG	2	21702155 - 21702552
FAN-SCF782	19	15.1	AAGAAAAGACCCACCCCACA	ATGGAATCAAAACACTGTGTTCGT	2	21709041 - 21709329
FAN-CON1144	19	16.9	AGCCATGAGTTCTCAATCACTGT	CACGTAAAACCCCGAAACCC	2	21747548 - 21747947
FAN-CON984	20	0.0	AGGAATCATACATTAGTTTCTCCT	TGCTTAGGGAACTGCAAATCCT	3	5608121 - 5608363
FAN-SCF55	20	0.6	AGGGTTGGCCTTGAAATCGA	GACCAGTGCGAACCTCTTGA	3	5867946 - 5868345
FAN-SCF11	20	16.1	GCTTCCAGAACCCTGAAGCT	AGTCAGGCTGTTGGGTAAGG	7	14002188 - 14002310
FAN-SCF625	21	0.0	ACGAAATACTGGCTCTGCTAGG	ACCCCAACAATTCCAAGCCA	7	8259563 - 8259843
FAN-CON1032	21	4.2	TCGTTAAGTGTATGATTCTCTCCA	GTGTGTTAAATTTATGTCGTACCA	7	6515448 - 6515725
FAN-SCF14	21	8.4	CCTCCCTAGCCTCCATCTCA	CTCGATCATCACCTTGGTCCA	2	11289091 - 11289202
FAN-SCF871	22	0.0	TGTGTGACTTTCTGTATGTTCA	CCAGGAACTTGTTGAGGAAAGC	3	9173551 - 9173871
FAN-SCF767	22	3.0	ACATGTTTCTGTGTCAGTTGGT	CCCTTCCGCTGATCTCTGTC	3	7351198 - 7351590
FAN-SCF97	22	6.5	TCCTTTGACATGATTGCCACG	AGTCAATGTCAACCTCCACTGG	3	6958967 - 6959366
FAN-CON1118	22	8.3	GCATTTGCTTTCAAGAGCGC	GCGTAAGTTTGTCCTGCAGG	3	7052910 - 7053309
FAN-CON1141	23	0.0	ACGATTCGACGCCTTTCGAT	ATTGGAGGAGGAGCATGCTT	3	15482868 - 15483267
FAN-CON1063	23	2.2	TCAGAAGAAATATAGCGTGGGT	AAGAATCCCACTCCAACAGCT	3	22688348 - 22688574
FAN-SCF587	23	3.5	GTCTGCAAAAGCAGGCAAGT	AGTCGTTGGATTTTTCTTTTAGGA	3	31471188 - 31471587
FAN-SCF915	23	3.5	CGCGCCAATTGATGAAGGAG	TCGAGCCTGTCATGGTCATG	3	31438822 - 31439221
FAN-SCF793	23	5.7	CCCACCCGAGCCCTTAAAAT	CGTCTGTGCCTATGTCGAGT	3	23487528 - 23487821
FAN-CON1085	23	6.8	GGACCTTGCCTTCCATGGAA	GGTTCTGAAACAAGATCCCCG	3	15926347 - 15926746
FAN-SCF287	23	7.4	TGCATGTGCCGTTTTCACAG	ACCAATGAAACCACGAACCT	3	16140230 - 16140639
FAN-SCF924	23	8.0	TCTGTCGGACAGTCTAAATGG	TGAACCCCTTCTACTGCAAAACT	3	16134694 - 16135091
FAN-SCF78	24	0.0	GACCATTCGTCGCGGATAGT	ACAGCACGTTCTGAGTTGAT	7	23293212 - 23293479
FAN-CON1025	24	6.8	TGAACCGGAAACTCTTACGT	TGGACCAAATCAAAACACACA	7	21504945 - 21505040
FAN-CON1154	25	0.0	GCTGAGTCCCTCGCATGTTA	ACATCACTTCAGCTGCACCA	5	16151073 - 16151358
FAN-CON1130	25	1.2	GCAGGTGGATGTCCGATTCA	GATTTGACGATCGGCCAAGC	5	16155482 - 16155766
FAN-SCF593	25	2.4	CCACTGCTGGAAGCCGAAAC	AGGATGGAATTTATCGCCCGA	4	7038979 - 7039083
FAN-SCF252	25	6.1	ACATGTCAAACGGTGAAAGCA	TATGCGAGCGAGCTAGATGC	3	24661969 - 24662295
FAN-SCF10	26	0.0	GGGCATTGTGGGTGCAAATA	CGGTGATCAAATTTGGGCCA	2	10258308 - 10258711
FAN-SCF458	26	4.2	ACCTAGTTTGAGCTATCTGGCA	CGTCCCACAAGTAGCTAGCT	2	3725132 - 3725214
FAN-SCF126	27	0.0	TCAACAGTTCAACACACTGA	TGTAAGCACTAGAACACTGGA	4	1265833 - 1266095
FAN-SCF164	27	0.0	TGCAGGCATTGGATCGTCAT	TCATTTGCAATGAAGCATCGCT	4	1524501 - 1524913
FAN-SCF390	27	2.4	TCTAGGGTTTGAGGACGCAA	GCATACAATTGGCACGTCCG	4	1770856 - 1771285
FAN-SCF383	27	3.1	CCGTCTCCCATCACTCCTCT	GGCGACTCAAGAGTTACCGA	3	11683723 - 11683764
FAN-SCF520	28	0.0	TGAGGATGTGCAACGTCTCA	TCGGGTTTCCAGATGTCGAC	2	19491481 - 19491897
FAN-SCF896	28	2.5	ATCTGAAGAGGCAAGCAGGG	CCAACTGATCTGCTCCTCCG	2	21512966 - 21513328
FAN-SCF76	29	0.0	GGATGAGAGGAACCGAGGGA	TTGAGATCTCGCACCGTTCC	7	7221386 - 7221749
FAN-SCF743	29	1.2	GCTCCAGCTTAGTCCATGCA	GCCTGAAGAAACACACACCG	4	15153915 - 15154307
FAN-CON1082	30	0.0	TCCGTCCATTGTGAAGAGGC	AGATCAACTTCAGAATCTGAGAA	3	196716 - 196972
FAN-SCF259	30	1.1	TGGGGTTGCACTTGAACAGT	GCGTTTCTTGCTTCAACGGT	3	117480 - 117879

Table 2

Statistics of raw reads, trimmed reads, and mapped reads of next generation resequencing data.

Plant material	Use	Raw reads	Trimmed reads			Mapped reads

		Total length (bp)	Total length (bp)	Trimmed/ Raw	Genome coverage	The number of mapped reads (%)	Mapped region of reference genome (bp)	Reference genome coverage

‘Senga-Sengana’	Male	30,682,186,222	21,846,702,806	71.2%	≒	193,915,797	530,090,126	76.0%
‘Senga-Sengana’	parent	30,682,186,222	21,846,702,806	71.2%	30.34X	(75.9%)	530,090,126	76.0%

‘Sulhyang’	Female	25,256,529,246	19,080,792,008	75.5%	≒	169,416,290	533,936,123	76.5%
‘Sulhyang’	parent	25,256,529,246	19,080,792,008	75.5%	26.50X	(77.0%)	533,936,123	76.5%

Table 3

Summary of single nucleotide polymorphism (SNP) detection and high resolution melting (HRM) primer design between strawberry cultivars ‘Sulhyang’ and ‘Senga-sengana.’

SNP type		The number of polymorphic SNP	The number of non-paralogous SNP	The number of HRM candidates	The number of HRM primer sets designed

‘Senga-sengana’	‘Sulhyang’				Total	Intergenic region	Genic region

							CDSz	Intron	Exon

Homozygous	Homozygous	206,019	8,128	6,750	2,324	1,212	504	416	192
Homozygous	Heterozygous	53,933	2,896	2,347	652	341	134	112	65
Heterozygous	Homozygous	74,367	4,220	3,460	1,338	676	298	247	117
Heterozygous	Heterozygous	87,161	5,604	4,460	1,154	443	458	181	72

Total		421,480	20,848	17,017	5,468	2,672	1,394	956	446

^zCDS indicates a coding sequence region except intron and exon regions.

Table 1 List of HRM primer sets used in this study.

Table 2 Statistics of raw reads, trimmed reads, and mapped reads of next generation resequencing data.

Table 3 Summary of single nucleotide polymorphism (SNP) detection and high resolution melting (HRM) primer design between strawberry cultivars ‘Sulhyang’ and ‘Senga-sengana.’
z

CDS indicates a coding sequence region except intron and exon regions.

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SNP 분자표지를 이용한 8배체 딸기( × Duch.) 유전자지도 작성

A genetic linkage map of allo-octoploid strawberry (Fragaria × ananassa Duch.) using SNP markers

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