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1、棉花學報CottonScience2016,28(2):129―137棕色棉莖尖基因槍遺傳轉化*侯云鵬袁馮鴻杰袁程文瀚袁張新宇袁孫杰袁劉永昌(石河子大學農學院/新疆兵團綠洲生態(tài)農業(yè)重點實驗室袁新疆石河子832003)摘要院為了建立高效的棉花莖尖遺傳轉化體系袁以棕色棉品種新彩棉5號為材料袁采用莖尖培養(yǎng)和基因槍遺傳轉化方法袁通過對凝固劑種類堯活性炭含量堯卡那霉素濃度堯滲透處理時間堯轟擊壓力堯轟擊距離等因素的優(yōu)化袁建立了棕色棉新彩棉5號莖尖基因槍遺傳轉化技術體系遙結果表明袁不同凝固劑種類堯活性炭含量堯卡
2、那霉素濃度堯滲透處理時間堯轟擊壓力和轟擊距離對棕色棉莖尖轉化率有明顯影響遙最佳的遺傳轉化體系為院前滲處理4h袁轟擊壓力7.586MPa淵1100psi冤袁轟擊距離9cm袁轟擊后恢復培養(yǎng)4d袁然后在MSB+8g窯L－1瓊脂+125mg窯L－1卡那霉素篩選培養(yǎng)基中篩選25d遙獲得抗性芽后袁在添加1g窯L－1活性炭的生根培養(yǎng)基中誘導抗性芽生根袁并獲得抗性再生植株遙通過PCR和qPCR檢測袁5200個莖尖共獲得16株陽性轉基因植株袁轉化率為0.31%遙該技術體系的建立為棕色棉的遺傳轉化奠定了基礎袁為彩色棉
3、育種提供了新的材料遙關鍵詞院棕色棉曰莖尖曰基因槍遺傳轉化曰轉化率中圖分類號院S562.03文獻標志碼院A文章編號院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窯L－1agarpowderand125mg窯L－1kanamycinandthenscreenedfor25dtoobtaintheresistantgerms;(4)then,1g窯L－1activatedcharcoalwasaddedtotheinductionmediumtoinducerootingoftheresistantgermstoobtainresistantregener
8、atingplants.AfterPCRandqPCRdetections,16plantspositiveforthetransgenewereobtainedfrom5200shoottips,withatransformationrateof0.31%.Theestablishmentofthistechnologysystemlaysafoundationforthegenetictransformationofbrowncottonandprovidesnewmateri
