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

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인삼 유전자원 장기보존을 위한 종자 초저온보존 조건 구명

Response of Pre and Post Treatments for Cryopreservation of Korean Ginseng Seeds on Recovering Viability

Korean Journal of Breeding Science 2014;46(4):408-416.
Published online: October 31, 2014

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*Corresponding Author : (youngyi@korea.kr, Tel: +82-31- 299-1804, Fax: +82-31-299-1894)
• Received: November 26, 2014   • Revised: December 17, 2014   • Accepted: December 17, 2014

© Tshe Korean Society of Breeding Science All rights reserved

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  • We investigated the effect of pre and post-cryopreservation treatment on the dehisced Ginseng (Panax ginseng C.A. Meyer) seeds germination rate. All seeds covered with endocarp and 96.3% of them were opened, initial moisture content (IMC) and germination rate were 55.9% and 84.0%, respectively. According to the pre and post-cryopreservation methods, highest seeds germination rate (GR) was observed in non-precooled seeds which recovered at 40°C, in which MC was 8-12% and desiccated at 15°C. However, by prolonged drying the seeds GR comparatively decreased under MC of below 2.2%, reaching below rate 4% regardless of pre and post- cryo treatments, whereas the seeds dried at 25°C airflow cabinet saved significantly high rate. The effects of desiccation and of cryopreservation on the survival and developmental pattern of roots and stems showed that the earliest appearance of the root in control seeds was detected after 10 days of bedding in GA medium, and the stem started to develop later on the third week of germination test. And, on the 30th day of the investigation stem growth rate significantly increased and showed equivalent data with root rate. Almost the same pattern showed the seeds after cryopreservation, but it significantly delayed in the development compared to control, where term of the investigation continued till 40 day than control 30 day. Additionally, non-precooling seeds in both desiccation and recovery methods were showed high root and stem growth compared to precooled treatments.
인삼은(Panax ginseng C.A. Meyer) 세계적으로 가장 약 효가 널리 알려진 약초 중 하나로서 건강 및 신체활력 뿐만 아니라 수명연장 기능 또한 보고되고 있는 기능성 유전자원 이다(Coleman et al. 2003, Ellis & Reddy 2002). 인삼은 종 자로 번식 가능하지만 일정 기간동안의 저장기간이 경과하여 야만 발아가 이루어지고, 발아과정에서의 환경요인에 따라 발 아율이 감소하는 등의 편차를 보이는 특성을 지니고 있다 (Kim et al. 2008, Kwon et al. 2001, Lee et al. 1995, Lee et al. 2004, Yoon et al. 2005).
종자은행의 종자 저장 방법은 국제기준을 비롯하여 많은 문헌에서 언급되어 왔으며(Hong & Ellis 1996), 최근에는 종 자 저장수명을 연장하고 장기간 보존할 수 있는 프로토콜 개 발을 위한 여러 가지 시도들이 이루어지고 있다. 핵심요인은 종자 수분이라고 할수 있는데 종자를 일정 수준 이상으로 건 조시키는 것은 저장수명에 거의 영향을 주지 못하며(Dickie et al. 1995, Ellis & Hong, 2006, Nagel & Borner, 2009), 오히려 노화를 가속화시킬 수 있다는 것(Vertucci & Roos, 1990, Walters, 1998)이 여러 연구를 통해 증명되어 왔다. 한 편, 생물조직을 액체질소를 이용하여 극한 저온(-196°C)에 저 장하는 초저온보존 방법은 세계적으로 널리 이용되고 있으며 특히 유전자원 보존에 있어서는 자원의 genetic potential 손실 없이 중장기 보존이 가능하게 한다(Benson & Withers, 1998). 이러한 과정은 초저온보존된 조직이 저장기간동안 대 사과정을 멈추게 함으로써 이루어진다(Engelmann 2004, Fatima et al. 2009). 이러한 초저온보존 프로토콜은 주로 다 양한 종의 건조종자, somatic 및 zygotic embryo를 중심으로 확립되어 왔다(Fatima et al. 2009, Gray et al. 1993, Sakai et al. 1990, Yi et al. 2013).
난저장(recalcitrant) 식물조직의 경우 초저온 처리 후에도 수명이 유지될 수 있는 수분함량을 유지하면서 보존에 알맞 게 탈수시키는 것이 필요한데 매우 까다로운 기술이 요구되 는 부분이다. 또한 건조에 대한 민감성이 다양한 여러 종들의 초저온보존 기술 개발은 많은 노력과 시간이 필요한 연구분 야라고 할 수 있으며(Chandel et al. 1995, King & Roberts 1980, Pence 1992 1995, Pritchard & Prendergast 1986, Wesley-Smith et al. 1992), 종마다 보존대상 조직부위가 다 른 것 또한 이러한 이유 때문이다(Berjak et al. 1993, Farrant et al. 1986, Finch-Savage 1992, Hong & Ellis 1990, Tompsett & Pritchard 1993).
인삼 종자(Kim et al. 2008, Yoon et al. 2005) 및 배배양 을 이용한 합성종자(artificial seed)의 효율적인 초저온보존 기술(Choi et al. 1999, Choi & Jeong 2002)은 앞선 연구들 에서 보고된 바 있으며, 이들 종자는 건조처리와 냉동조건을 견딜 수 있다는 시험결과가 도출되기도 하였다. 본 연구의 목 적은 인삼 종자를 초저온보존하기 위한 몇가지 조건을 구명 하고자 하는 것으로서, 인삼 개갑종자의 초저온보존에 있어 사전 동결처리 유무 및 보존 후 해동단계의 조건에 따른 회복 발아율을 조사함으로써 보존효율을 알아내고자 하였다. 또한 종자수분함량 및 초저온보존 처리가 회복된 유묘의 뿌리 및 줄기 발달양상에 미치는 영향을 구명하고자 하였다.
종자시료
인삼(Panax ginseng C.A. Meyer) 종자는 국립원예특작과학 원 인삼특작부에서 2013년에 수확하여 개갑처리 한 후 90일간 2°C에서 층적처리하여 2014년 2월부터 국립농업과학원 농업유 전자원센터에서 시료로 이용하였다. 종자의 크기에 따른 발아 율 차이는 없다고 보고(Kim et al. 2014)된 바에 따라 크기에 상관없이 임의 선발하여 시험에 사용하였으며 개갑율은 93.6%, 초기 수분함량(IMC)은 55.9%, 초기 발아율은 84.0% 이었다.
수분함량 측정 및 건조처리
종자 수분함량(MC)은 105°C 오븐에서 24시간 건조하면서 종자 무게를 조사함으로써 산출하였다. 종자가 일정수분함량 (DMC)에 이르렀는지를 조사하기 위하여 건조 전후의 종자무 게를 이용한 다음의 식을 적용하였다.
Weight of seed (g) at DMC% =(100-initial MC%) initial seed weight(g)100 - DMC%
종자 건조율은 종자량(종자층의 깊이), 건조상 내부의 공기 유동 및 종에 따라 다르다. 본 연구에서는 종자 건조처리를 두가지 방법으로 실시하였다. 첫 번째는 종자를 15°C, RH 10-12% 의 일반적인 종자건조실에 처리하였고, 두 번째는 2 5°C, RH22-23%로 조절된 airflow 챔버에 처리하였다. 4-5% 미만의 종자수분함량을 설정하기 위해서는 추가적으로 40% 오븐에서 1시간 건조처리 하였으며 모든 시험은 100립씩 3반 복으로 시행하였다.
초저온보존, 해동 및 종자발아
종자수분함량 설정 후(2-12%) 각각 5 mL cryovial에 50립 씩 담아 액체질소에 사전 동결단계 없이 바로 담그고, 비교구 로서 같은 한 세트는 보존 전 동결단계를 거치도록 하였다. 0 ± 0.3°C까지 서서히 온도가 저감되도록 프로그래밍된 챔버에 서 24시간 처리한 후 3시간동안 –1°C까지 저감되도록 하고 이후 1시간 간격으로 –24°C까지 저감시킨 후 14시간동안 저 장하였다. 액체질소 처리는 30일간 지속하였다.
액체질소 저장 후 해동은 40°C water bath에서 2시간동안 처리한 구와 4°C 챔버에서 24시간 상치한 후 실온에 30분 처 리한 구를 비교하였다.
발아 조사를 위하여 1% NaOH 및 70% 에탄올로 소독한 종자를 증류수로 수세한 후 25°C에서 3시간 건조시켜 200 mg/L-1GA3 용액으로 적신 paper에 Between Paper법으로 치 상하고 10°C 챔버에서 배양하였다(Kim et al. 2008, Rajametov et al. 2014, Yoon et al. 2005). 발아 조사는 10일 간격으로 50일간 실시하였으며 3 mm 이상 유근이 출현한 경우를 정상 종자로 판정하였다(Kim et al 2008). 초저온보존을 위한 사전 동결처리 및 사후 해동처리 및 유묘생육 영향을 조사하기 위 하여 종자에서 출현된 뿌리 및 자엽 생육을 조사하였다.
종자 건조조건
인삼 개갑종자의 수분함량은 15°C에서보다 25°C에서 빠른 속도로 감소하여 5일만에 임계점까지 떨어졌으나, 15°C에서 는 같은 수준으로 떨어지기까지 40일이 소요되었다(Fig. 1).
Fig. 1.
Effects of the drying methods on the seed desiccation rate.
KJBS-46-408_F1.gif
발아율 변화
종자수분함량 및 초저온보존 전후 처리에 의해 발아율은 Fig. 2와 같이 변화를 보였다. 종자수분 15°C에서 종자수분 8-12%로 건조된 종자가 사전 동결단계를 거치지 않고 초저 온보존된 후 40°C에서 2분간 해동처리된 되었을 때 가장 높 은 발아율을 보였다. 같은 수분함량의 대조구는 유의하지는 않으나 약간 더 높은 발아율을 나타냈다.
Fig. 2.
Effects of desiccation and recovery conditions on the germination rate of cryopreserved dehisced Ginseng seeds under different moisture content (FW) . Coated seeds were stored in LN for 30 day. The vertical bars indicate SD (n= 3).
KJBS-46-408_F2.gif
특히 초저온보존을 위한 사전 동결처리를 한 모든 종자들 은 처리하지 않은 종자들보다 모두 발아율이 낮은 것으로 조 사되었다. 또한 15°C에서 건조된 처리구는 해동조건과 무관 하게 수분함량 5% 이하로 떨어지면 급속히 발아율이 감소하 는 것으로 나타나 25°C 건조 처리구보다 저감율이 빠른 것으 로 나타났다.
유근 및 자엽발달에 미치는 영향
종자 건조 및 저장방법에 따라 식물체의 생장은 달라질 수 있으므로 본 연구에서는 15°C에서 수분함량 8.8%로 조절된 종자와 25°C에서 종자수분 9.6%로 조절된 종자를 발아시켜 발아 기간동안 유근과 자엽생장을 조사하였다. 그 결과 유근 과 자엽 생장은 건조 방법과 상관없이 사전 동결처리 없이 보 존 후 40°C에서 2분간 해동처리한 구에서 가장 높은 것으로 조사되었다(Fig. 3, 4). 그러나 초저온보존 후 유근 및 자엽의 생장은 초저온처리 하지 않은 일반 종자(IMC 55.9%)보다 현 저히 낮은 것으로 나타났다.
Fig. 3.
Effects of desiccation and pre and post cryopreservation treatments on the growth ability of root and stem. The vertical bars indicate SD (n= 3).
KJBS-46-408_F3.gif
Fig. 4.
Growth rate of Ginseng seeds hypocotyl (a fused hypocotyl and taproot) depends on pre and post cryopreservation treatments (30 days of culture).
KJBS-46-408_F4.gif
대조구에서의 유근 출현은 GA 배지에 치상한지 10일 이내 에 관찰되었으며 자엽은 발아한지 3주 후에 발달되기 시작하 였다. 30일 동안 자엽생장은 유근생장과 함께 증가세를 나타 냈다.
초저온보존 후, 30일 동안 유근 및 자엽 발달은 대조구보다 현저히 감소하는 것으로 나타났으나 40일째 조사결과는 이후 식물체의 생장율이 다른 양상을 나타낼 수도 있음을 시사하 였다(Fig. 3, 대조구 종자발아 종료로 데이터 없음).
생물 조직의 효율적인 초저온보존을 위해 많은 프로토콜들 이 개발되고 있다. 초저온보존은 식물 유전자원의 안정적이고 비용 효율적인 장기보존을 위해 유용하게 활용될수 있기 때 문이다. 식물 종 각각에 적합한 프로토콜 개발을 위해 다양한 연구들이 시도되어 왔다. 보존 전처리 뿐만 아니라 해동, 발아 배지 등과 같은 보존 후처리 또한 장기 보존에 있어 중요한 요인으로서 종자 생육과 활력에 크게 영향을 주는 것으로 알 려지고 있다.
극한 저온에서 일정 식물조직을 장기간동안 보존하기 위해 서는 수분이 가장 중요한 요인으로 작용하는데, 이는 초저온 보존된 세포의 결빙(ice crystallization) 유도로 인한 피해의 원인이기 때문이다(Ashworth 1992, Kim et al. 2008, Sparks et al. 2001, Yoon et al. 2005). 뿐만 아니라 수분함량은 배 생장, 호르몬 농도, 화학조성 및 대사활성과 같은 생물·생리적 상태에 중요한 역할을 하므로 수분함량은 해당 조직의 장기 보존에 있어 가장 중요한 결정 요소라고 할 수 있다(Kim et al. 2008, Kim et al. 2014).
최근 Kim 등(2014)은 몇몇 인삼 종자의 내생 GA 호르몬 (GA1-34)이 배 생장율을 현저히 증가시킨다고 보고하였고, 발 아기간 단축 및 휴면 타파를 목적으로 GA3가 주로 사용되고 있으나, GA4가 가장 효과적이라는 연구도 발표된 바 있다(Kim et al. 2008, Rajametov et al. 2014, Yoon et al. 2005).
본 연구에서는 인삼 개갑종자의 수분함량이 25°C air flow 챔버에서 15°C 건조실에서 보다 빠르게 감소하였고, 종자수 분 임계점까지 이르는 시간도 각각 5일, 30일로 나타났다. 즉, 종자 수분을 10%로 조절하기 위해서 25°C 챔버에서 3 시간, 15°C 건조실에서 5일이 소요되는 것으로 조사되었다. 이는 생육상 및 건조실에서 인삼종자를 건조처리 및 비교한 다른 연구 결과들과도 유사한 양상을 보였다(Kim et al. 2008, Yoon et al. 2005). Kim 등(2008)은 건조 및 흡습 방법을 달 리 하여도 건조 종자의 발아율에는 영향이 없다고 보고한 바 있으나 처리된 온도 및 상대습도 조건에 대한 정보는 기술되 어 있지 않다. 본 연구에서는 다양한 식물종을 서서히 건조시 키는데 사용되는 15°C 건조실(Hong & Ellis 1996, Hong & Ellis 2006)을 이용하여 종자수분이 4-5%로 떨어지는데에 40 일이 소요되었다.
특정 종의 식물은 종자수분이 일정 수준 이하로 감소하면 종자 활력 및 생육이 불가능해 진다는 것은 잘 알려진 사실이 다(Vertucci et al. 1990). 또한 일정 수준의 수분함량은 식물 조직의 저장 수명을 결정하는데, 이때의 수분함량 수준은 식 물 종마다의 ‘안전 최소수분(LSMC)’ 및 ‘어는점을 견딜 수 있는 최대수분(HMFL)’이 고려되어야 한다. 건조된 일반종자 는 액체질소 처리시 2-28%의 수분함량 조건에서(유지 종자 의 경우 3% 전후, 전분 종자의 경우 4-6%) 생존 가능한 것으 로 알려져 있다(Chin & Krishnapillay 1989, Das Bhowmik et al. 2011, Fassil & Engels 1997, Hong & Ellis 1996, Stanwood 1985, Yoon et al. 2005).
Yoon 등(2005) 및 Kim 등(2008)에 의하면 인삼 종자의 LSMC는 7%, HMFL은 12-13% 로서 인삼 종자수분이 13.5% 이상이면 결빙현상에 의해 결과적으로 탄수화물에 비해 지질 과 단백질의 비율이 높아지는 현상을 초래하게 된다. 따라서 액체질소 저장에 의한 종자 피해를 방지하기 위해 본 연구에 서는 종자수분은 2-12%로 설정하였다. 이 경우 높은 발아율 을 보인 종자의 수분함량은 HMFL에 가까웠고, 60% 이하의 낮은 발아율을 보인 종자의 수분함량은 LSMC에 가까웠다. 본 연구에서는 15°C에서 건조된 종자시료에서 HMFL은 약 12%로 조사되었고, LSMC는 7% 이상으로 가장 낮은 발아 율을 보여 선행 연구들과 유사한 결과를 보였다. 반면 25°C에 서 건조된 종자는 다소 다른 양상을 보였다.
액체질소에 저장된 개갑종자의 발아율은 보존 전후처리에 따라 다른 결과를 나타냈다. 가장 높게 나타난 발아율은 80.7% 로 수분함량이 12.4%이고 사전처리 없이 보존 후 40°C에서 해동처리한 종자에서 관찰되었다. 그러나 종자 수분이 2.2% 이하로 감소하면 발아율은 감소하였고 4% 이하가 되면 초저 온보존 전후 처리효과는 영향이 없게 되는 것으로 나타났다.
25°C에서 수분함량 3%로 건조된 종자는 15°C에서 건조된 종자의 같은 수분함량 수준에서보다 높은 발아율을 나타냈는 데 이는 온도, 상대습도와 같은 건조 조건이 발아율 뿐만 아 니라 관련된 호흡율, 대사 및 수명과 밀접한 관계가 있음을 시사한다(Lasseran 1988, Leon-Lobos et al. 2005, Miranda et al. 2010, Sanderson et al. 1989, Vertucci 1989). 일정 범 위의 종자 수분은 산소 흡수, 단백질 혹은 생물학적 조직 변 화에 영향을 주고(Leopold & Vertucci 1989, Senaratna et al. 1986, Vertucci et al. 1990), 더 나아가서는 세포 조직의 구조적, 생리적 변화를 초래할 수 있다(Wesley-Smith et al. 2001).
Vertucci(1989)Leopold와 Vertucci(1989)는 산소가 빠르게 소모되기 시작하는 수분함량은 ATP 합성, 호흡억제 와 같이 미토콘드리아의 전자공여가 일어날 수 있는 수분함 량이라고 하였다. 탈수 과정이 일어나는 동안 발아율 감소로 인한 생물학적 기작들 중에서 세포막 기능의 손상은 전도도, 지 질 과산화 및 용출전해질, 종자 탄수화물 등으로 측정될 수 있 다(Chandel et al. 1995, Crowe et al. 1984, Roberts 1979, Senaratna & McKersie 1986). 본 연구에서 15°C에서 건조 된 수분함량 7% 이하의 인삼 개갑종자는 상대적으로 발아율 이 낮았는데 이는 상대적으로 긴 건조기간동안 탈수과정이 지속되면서 액포의 민감도 증가에 의해 종자 노화 및 종피 손 상, 세포의 기계적 상해 등이 유도됨으로써 나타난 결과일 가 능성도 고려해야 한다(Wesley-Smith et al. 2001). 가장 낮은 발아율은 25°C에서 수분함량 11%로 건도된 종자에서 조사되 었는데, 이는 짧은 건조 기간동안 갑작스런 대사속도의 변화 로 호흡 및 수분 손실이 증가하여 세포 내외부 전반적인 수분 함량 불균형에 기인한 것으로 생각될 수 있다. 수분함량이 완 전한 hydration과 기능장애로 인한 유해반응을 줄이는 대사를 조절할 수 있을 때 정상적인 기능은 이루어진다(Berjak & Pammenter 1997 and 1999, Leprince et al. 2000, Pammenter et al. 1991, Walters et al. 2001).
앞서 언급한 바와 같이 전처리 없이 액체질소에 보존 후 4 0°C에서 빠르게 해동한 종자가 유의하게 높은 발아율을 나타 냈으나, 보존 전 동결처리구는 유근과 자엽 생장율이 상대적 으로 높게 나타났다. 시각적으로 유근 및 자엽 생장은 건조방 법 및 종자수분 함량과는 무관하게 보였으나(Fig. 4) 종자수 분 함량이 감소함에 따라 유근과 자엽 생장율은 유의하게 감 소하였고 발달이 저해되었으며 이 때의 종자는 15°C에서 수 분함량 5% 이하로 건조된 상태였다.
본 연구 결과, 인삼 개갑종자의 초저온보존 최적 조건은 종 자를 전처리 없이 바로 액체질소 보존하고 40°C 해동처리 하 는 것이다. 그러나 종자의 수분함량과 세포기능 활성은 유근 과 자엽 발달단계에 유의한 영향을 줄 수 있고 초저온보존시 최적 종자수분은 식물조직의 생리적 단계에 따라 다르게 나 타날 수 있다. 종자를 전처리 없이 액체질소에 담갔을 때 생 체 활성은 일시적으로 정지함으로써 내부 결빙 형성이 사전 동결처리한 종자보다 유의하게 적게 이루어져 사전 동결처리 가 초저온에 대한 순화효과 보다는 오히려 저온스트레스 요 인으로 작용한 것으로 생각된다. 수분함량 13% 이상의 인삼 종자가 동결이나 해동될 때, 지질의 결빙(응고)이 일어난다고 보고된 바도 있으며(Kim et al. 2008), 이는 세포 내외에 불 규칙한 모양의 ice를 생성함으로써 결과적으로 세포벽에 상해 를 초래한다고 알려져 있다(Leprince et al. 2000, Rajashekar & Burke 1996). 또한 즉각적인 동결 및 해동은 극저온쇼크 와 같은 스트레스와 일시적인 생체활성 변환을 최소화하는데 유효할 것으로 생각된다.
결론적으로 인삼 개갑종자는 고유의 건조 내성을 갖고 있 으며, 수분함량과 극저온에 대한 민감도가 발아율과 유근 및 자엽 생장에 유효한 영향을 미친다. 종자수분함량이 8-12%인 인삼 종자는 액체질소 내에 잘 저장될 수 있으며 40°C에서 해동처리하면 보존 후 발아에 효과적이다.
인삼 개갑종자의 초저온보존 효과 및 최적조건을 구명하기 위하여 건조 조건과 수분함량 및 액체질소 보존 전 동결처리 와 보존 후 해동처리에 따른 초저온보존 효과를 조사하였다. 건조처리는 25°C air flow 챔버(RH 10~12%)와 15°C 건조 실(RH 22~25%)을 이용하여 실시하였고 보존 전 동결은 – 24°C에서 30일간 처리하였으며 보존 후 해동은 4°C에서 24 시간, 40°C에서 2분 처리구를 각각 비교하였다. 그 결과 15°C 에서 종자수분 8-12%로 건조된 종자가 보존 전 처리 없이 액 체질소에 보존된 후 40°C에서 해동되었을 때 발아율이 가장 높은 것으로 나타났다. 그러나 종자수분이 2.2% 이하일 때 건조가 지속되면 발아율은 감소하는 것으로 나타났으며, 4% 이하에 이르면 보존 전후 처리효과는 나타나지 않는 것으로 조사되었다. 반면 25°C에서 건조된 종자는 상대적으로 높은 발아율을 유지하였다. 건조 및 초저온보존에 의한 인삼 개갑 종자의 유근 및 자엽 생장율을 조사한 결과, GA배지 치상 10 일 후 무처리한 대조구에서 첫 유근이 출현하였으며 3주 후에 는 자엽발달이 시작되었다. 30일째에는 자엽생장율이 유근발 달과 함께 유의하게 증가하는 것으로 나타났다. 치상 30일까 지 초저온보존에 의한 생장율은 같은 양상을 보이면서 대조 구보다 현저히 낮게 나타났으나 40일째에는 양상은 같았으나 회복율이 대조구보다 증가세를 보였다. 또한 보존 전 동결처 리 없었던 종자는 동결처리한 종자보다 모든 처리구에서 높 은 유근 및 자엽 생장율을 보였다.
본 연구는 농촌진흥청 연구사업(과제명: 종자 유전자원 안 전보존 및 품질관리체계 구축 연구, 과제번호: PJ008553)의 지원으로 이루어진 것임.
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Response of Pre and Post Treatments for Cryopreservation of Korean Ginseng Seeds on Recovering Viability
Korean. J. Breed. Sci.. 2014;46(4):408-416.   Published online December 31, 2014
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Response of Pre and Post Treatments for Cryopreservation of Korean Ginseng Seeds on Recovering Viability
Korean. J. Breed. Sci.. 2014;46(4):408-416.   Published online December 31, 2014
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Response of Pre and Post Treatments for Cryopreservation of Korean Ginseng Seeds on Recovering Viability
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Fig. 1. Effects of the drying methods on the seed desiccation rate.
Fig. 2. Effects of desiccation and recovery conditions on the germination rate of cryopreserved dehisced Ginseng seeds under different moisture content (FW) . Coated seeds were stored in LN for 30 day. The vertical bars indicate SD (n= 3).
Fig. 3. Effects of desiccation and pre and post cryopreservation treatments on the growth ability of root and stem. The vertical bars indicate SD (n= 3).
Fig. 4. Growth rate of Ginseng seeds hypocotyl (a fused hypocotyl and taproot) depends on pre and post cryopreservation treatments (30 days of culture).
Response of Pre and Post Treatments for Cryopreservation of Korean Ginseng Seeds on Recovering Viability