生姜脱毒快繁和病毒检测技术研究.docVIP

生姜脱毒快繁和病毒检测技术研究 　　摘要 本文以感染TMV和CMV的湘西本地小黄姜为试材，采用热处理、茎尖培养结合添加病毒唑的方法进行脱毒处理以获得无病毒的优质生姜种苗，运用RT-PCR方法检测病毒，以期探讨出生姜最适宜的脱毒快繁及病毒检测技术。结果表明，将生姜块茎置于40 ℃条件下催芽，显微镜下剥取0.2～0.5 mm的茎尖进行培养，最佳诱导培养基的配方为MS+6-BA 2 mg/L+NAA 0.5 mg/L+病毒唑40 mg/L，继代培养基中将MS培养基中的蔗糖浓度调整为80 g/L，即可实现生姜试管苗的增殖、生根及壮苗的一步培养，培养周期以30～40 d为宜。建立了一套生姜CMV和TMV 2种病毒的RT-PCR检测体系，运用该检测方法对生姜脱毒试管苗进行检测，病毒检出率为0，生姜脱毒率达100%。 　　关键词 生姜；茎尖脱毒；同步培养；病毒唑；病毒检测 　　中图分类号 S632.5 文献标识码 A 文章编号 1007-5739（2017）22-0049-03 　　Abstract In order to obtain high quality ginger seedlings without virus，the yellow ginger of Xiangxi three main virus detoxification methods were studied.Taking TMV and CMV infection Yellow ginger of Xiangxi as experimental materials，the heat treatment，stem tip cultivation method of adding ribavirin combination，in the process of test using RT-PCR method to detect virus，the most suitable detoxification and virus detection technology of ginger were discussed.The results showed that while putting the ginger tuber for germination at 40 ℃，and then wring 0.2-0.5 mm size stem tip under the microscope for culturing.The optimal medium for induction was MS+6-BA 2 mg/L+NAA 0.5 mg/L+virazole 40 mg/L.In the subculture medium，when the sucrose concentration in the MS medium was adjusted to 80 g/L，the ginger test tube′s proliferation，taking root and strong seedlings can be realized by one step，and the suitable culture cycle is 30-40 days.A RT-PCR detection system for CMV and TMV viruses was established.The virus detection rate of ginger detoxification test tube was zero by RT-PCR，ginger detoxication rate is 100%. 　　Key words ginger；stem tip detoxification；synchronization culture；ribavirin；virus detection 　　生姜?榻?科姜属多年生宿根植物，营养丰富，是药、食、加工等多用途经济作物，是我国重要的外贸产品。湘西自治州地处我国生姜主产区，生姜种植历史悠久，全州生姜种植面积约3 333.33 hm2 [1]。生姜在生产上长期采用无性繁殖，重茬种植，易感染多种病毒，使姜体内积累多种病毒，从而导致生姜产量下降、品质降低，种性退化，严重制约了生姜的生产发展。国内外目前尚无高抗病毒的生姜品种和高效杀病毒药剂，因此，亟需通过组织培养手段对现有优良品种进行脱毒和复壮，提高品种自身抗逆性和增产潜力。研究表明[2-3]，侵染生姜的病毒主要为黄瓜花叶病毒（CMV）和烟草花叶病毒（TMV），病毒与姜竞争生长必需的营养成分，使生姜出现局部或系统花叶褪绿、植株矮化或叶畸形等症状，从而导致生姜产量降低 30%，严