甘肃农业大学学报
甘肅農業大學學報
감숙농업대학학보
JOURNAL OF GANSU AGRICULTURAL UNIVERSITY
2014年
3期
53-59
,共7页
二穗短柄草%HTA基因%载体构建%rat5
二穗短柄草%HTA基因%載體構建%rat5
이수단병초%HTA기인%재체구건%rat5
brochypodium distachyon%HTA gene%vector construction%rat5
通过PCR扩增,将二穗短柄草基因Bradi1g25390(BdHTA11)、Bradi1g25400(BdHTA12)与拟南芥基因At1g54640(AtHTA14)连接到植物表达载体 pBINPLUS 的多克隆位点,分别构成3个重组载体 pBINPLUS-BdHTA11、pBINPLUS-BdHTA12和 pBINPLUS-AtHTA14,载体 pBINPLUS 由烟草花叶病毒35s 启动子启动目的基因的表达。采用农杆菌侵染拟南芥花序的转化方法,将这3个基因转化到拟南芥 rat5突变体中,经过筛选获取转基因植株。经半定量 RT-PCR分析,初步确定目的基因已经整合到拟南芥 rat5突变体基因组中并过量表达。
通過PCR擴增,將二穗短柄草基因Bradi1g25390(BdHTA11)、Bradi1g25400(BdHTA12)與擬南芥基因At1g54640(AtHTA14)連接到植物錶達載體 pBINPLUS 的多剋隆位點,分彆構成3箇重組載體 pBINPLUS-BdHTA11、pBINPLUS-BdHTA12和 pBINPLUS-AtHTA14,載體 pBINPLUS 由煙草花葉病毒35s 啟動子啟動目的基因的錶達。採用農桿菌侵染擬南芥花序的轉化方法,將這3箇基因轉化到擬南芥 rat5突變體中,經過篩選穫取轉基因植株。經半定量 RT-PCR分析,初步確定目的基因已經整閤到擬南芥 rat5突變體基因組中併過量錶達。
통과PCR확증,장이수단병초기인Bradi1g25390(BdHTA11)、Bradi1g25400(BdHTA12)여의남개기인At1g54640(AtHTA14)련접도식물표체재체 pBINPLUS 적다극륭위점,분별구성3개중조재체 pBINPLUS-BdHTA11、pBINPLUS-BdHTA12화 pBINPLUS-AtHTA14,재체 pBINPLUS 유연초화협병독35s 계동자계동목적기인적표체。채용농간균침염의남개화서적전화방법,장저3개기인전화도의남개 rat5돌변체중,경과사선획취전기인식주。경반정량 RT-PCR분석,초보학정목적기인이경정합도의남개 rat5돌변체기인조중병과량표체。
The genes of Brachypodiumdistachyon Bd1g25390(BdHTA11),Bd1g25400(BdHTA12)and arabidopsis gene At1g54640(AtHTA14)connected to the multiple cloning site of PBINPLUS by PCR.This constituted three recombinant vectors-pBINPLUS-BdHTA11,pBINPLUS-BdHTA12 and pBINPLUS-AtHTA14.And this pBINPLUS was expressed by the tobacco mosaic virus 35s promoter to drive target genes.The three genes were transformed into Arabidopsis rat5 mutant by the technology of agrobacterium infecting Arabidopsis thaliana inflorescence.Transgenic plants were screened out,which was determined preliminarily that the target genes had been integrated into Arabidopsis rat5 mutant genome and overex-pressing by semi-quantitative PCR.
