半夏凝集素基因在水稻中的转化及筛选

Transformation of Pinellia ternata Agglutinin Gene into Rice and Preliminary Screening

为了获得转基因抗虫水稻,将半夏凝集素基因pta导入水稻,并进行鉴定和筛选。在抗虫基因pta的5'端加2×35S启动子,两端分别添加HindⅢ和Spe I酶切位点,并进行人工合成。将pta与植物表达载体p1301连接,经双酶切和测序鉴定。结果表明:重组表达载体p1301-pta构建成功;载体p1301-pta转化农杆菌LBA4404后用浸染法将pta基因导入水稻恢复系辐恢838中,对转化再生植株进行GUS染色以及GUS和pta基因的PCR鉴定后,共得到16株转基因阳性植株。

To obtain insect-resistant transgenic rice,P. ternate agglutinin（ pta） gene was transferred into rice,which was identified and screened. The Hind Ⅲ and Spe I enzyme cutting sites were separately added at both ends of the pta plus 2 × 35 S promoter. This sequence was artificially synthesized. Gene pta was ligated into vector p1301 by double enzyme digestion and DNA ligase to construct ecombinant expression vector p1301-pta. Identification of double enzyme digestion,and sequencing showed that the vectoe p1303-pta was successfully constructed. Then p1303-pta was transformed into Agrobacterium tumefaciens LBA4404 that infected calli of rice Fuhui 838 into which pta was transferred so as to make pta transfer into of rice restore by LBA4404. The transgenic rice plants were identified by GUS staining and PCR amplification for GUS and pta genes. A total of 16 positive transgenic rice plants were obtained.