拟南芥SSP1基因GFP载体构建及GFP植株筛选  

Construction of Arabidopsis SSP1-GFP Vector and Identification of Transgenic Lines

作　　者：吴席 樊婷婷 方雪 徐洁娜 曹树青 WU Xi;FAN Ting-ting;FANG Xue(School of Food Science and Engineering,Hefei University of Technology,Hefei,Anhui 230009)

机构地区：[1]合肥工业大学食品科学与工程学院,安徽合肥230009

出　　处：《安徽农业科学》2018年第21期119-121,共3页Journal of Anhui Agricultural Sciences

摘　　要：[目的]以野生型拟南芥为材料构建SSP1基因GFP载体及GFP转基因植株。[方法]通过提取野生型拟南芥的RNA,反转录成为c DNA,并以c DNA为模板,利用高保真酶,进行SSP1基因的克隆,将克隆所获得SSP1基因和p XB94-GFP质粒进行双酶切,利用T4-DNA ligase进行连接,并转化进大肠杆菌DH5α感受态细胞中,通过菌落PCR和测序获得阳性单克隆。通过质粒抽提试剂盒提出构建好的重组质粒,并将其转入农杆菌GV3101中,通过菌落PCR鉴定出阳性单菌落。将获得的阳性菌通过花序浸染法转入野生型拟南芥中,并通过抗性筛选获得阳性植株。[结果]克隆SSP1基因成功,菌落PCR显示SSP1基因成功连接到p XB94-GFP质粒上并成功转入农杆菌中,抗性筛选获得了SSP1-GFP转基因植株。[结论]成功获得SSP1-GFP转基因植株,为接下来进一步研究SSP1基因功能奠定了基础。[Objective]To build up the vector of Arabidopsis SSP1-GFP and transgenic lines.[Method]Reverse transcription to cDNA by extracting the RNA of wild type Arabidopsis.The CDS fragment of SSP1 gene was amplified by PCR.After treatment with restriction enzymes and T4-DNA ligase,the gene was connected to plasmid.Transfer the connection product to DH5αand a positive clone was identified.Then,transfer the recombinant to GV3101.The floral-dip method was used to transfer it into wild type Arabidopsis.Finally,we obtained positive plants through resistance screening.[Result]We succeeded in cloning SSP1 gene.The recombinant plasmid was obtained and we got the transgenic lines.[Conclusion]The successful acquisition of SSP1-GFP transgenic lines laid the foundation for further research on the function of SSP1 gene.

关 键 词：拟南芥 SSP1 载体 GFP转基因植株 

分 类 号：Q939.9[生物学—微生物学]