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1、棉花学报CottonScience2016,28(2):129―137棕色棉茎尖基因枪遗传转化*侯云鹏袁冯鸿杰袁程文瀚袁张新宇袁孙杰袁刘永昌(石河子大学农学院/新疆兵团绿洲生态农业重点实验室袁新疆石河子832003)摘要院为了建立高效的棉花茎尖遗传转化体系袁以棕色棉品种新彩棉5号为材料袁采用茎尖培养和基因枪遗传转化方法袁通过对凝固剂种类尧活性炭含量尧卡那霉素浓度尧渗透处理时间尧轰击压力尧轰击距离等因素的优化袁建立了棕色棉新彩棉5号茎尖基因枪遗传转化技术体系遥结果表明袁不同凝固剂种类尧活性炭含量尧卡
2、那霉素浓度尧渗透处理时间尧轰击压力和轰击距离对棕色棉茎尖转化率有明显影响遥最佳的遗传转化体系为院前渗处理4h袁轰击压力7.586MPa渊1100psi冤袁轰击距离9cm袁轰击后恢复培养4d袁然后在MSB+8g窑L－1琼脂+125mg窑L－1卡那霉素筛选培养基中筛选25d遥获得抗性芽后袁在添加1g窑L－1活性炭的生根培养基中诱导抗性芽生根袁并获得抗性再生植株遥通过PCR和qPCR检测袁5200个茎尖共获得16株阳性转基因植株袁转化率为0.31%遥该技术体系的建立为棕色棉的遗传转化奠定了基础袁为彩色棉
3、育种提供了新的材料遥关键词院棕色棉曰茎尖曰基因枪遗传转化曰转化率中图分类号院S562.03文献标志码院A文章编号院1002-7807(2016)02-0129-0910.11963/issn.1002-7807.201602005*HouYunpeng,FengHongjie,ChengWenhan,ZhangXinyu,SunJie,LiuYongchang(832000,冤Togeneticallytransformcottonshoottips,weestablishedabiolistic
4、genetictransformationsystemthoughtheplantshoottipculturetechniqueandthegenegun-mediatedtransformationmethod,usingtheshoottipsofbrowncottonXincaimianNo.5,aL.varietyculturedinXinjiangasmaterials.Severalcriticalfactorsofthesystemwereassessed,includingthe
5、typeofcoagulant,theactivatedcharcoalconcentration,thekanamycinconcentration,thepenetrationprocessingtime,andthebombardmentpressureanddistance.Resultsshowedthateachfactorclearlyinfluencedthetransformationrateofbrowncottonshoottips.Anoptimalbiolisticgen
6、etictransformationtechnologysystemwasdetermined:(1)shoottipswereculturedfor4hbeforebeingbombardedintheosmoticmedium;(2)theshoottipswerebombardedwithheliumpressureof7.586MPa(1100psi),andabombardmentdistanceof9cm;(3)afterculturingfor4dintherecoverymediu
7、m,shoottipsweremovedtoMSBscreeningmediumcontaining8g窑L－1agarpowderand125mg窑L－1kanamycinandthenscreenedfor25dtoobtaintheresistantgerms;(4)then,1g窑L－1activatedcharcoalwasaddedtotheinductionmediumtoinducerootingoftheresistantgermstoobtainresistantregener
8、atingplants.AfterPCRandqPCRdetections,16plantspositiveforthetransgenewereobtainedfrom5200shoottips,withatransformationrateof0.31%.Theestablishmentofthistechnologysystemlaysafoundationforthegenetictransformationofbrowncottonandprovidesnewmateri
