摘要
将水稻(Oryza sativa L.)10 kD富硫醇溶蛋白基因。cDNA(PLG)分别与Patatin Class I启动子、CaMV35S启动子和NOS 3＇终止子融合,构建了表达载体pBinLG、pBinLGP。表达载体通过直接法转入农杆菌(Agrobacterium tumefaciens)LBA4404(pAL4404),然后转化马铃薯(Solanum tuberosum L.)薯块,在含有100mg/L卡那霉素的抗性培养基上再生成植株。对抗性植株的NPTⅡ酶活性检测、总DNA的PCR扩增及Southern杂交、总RNA的点杂交、蛋白质Western杂交,证明目的基因已导入马铃薯细胞中,整合到马铃薯基因组上,井能正确地表达。
Using 10 kD sulfur-rich prolamin gene of rice (PLG) as target gene, the authers constructed the expression vectors pBinLG and pBinLGP, which contained CaMV 35S promoter/PLG/ NOS terminator, and Patatin Class Ⅰ promoter/PLG/NOS terminator respectively. They were transformed into Agrobacterium tumefaciens strain LBA4404 (pAL4404) by direct transformation method with incubating the leaf and tuber explants of potato ( Solanum tuberosum L.) with LBA4404 (pAL4404) and selecting in the medium containing 100 mg/L kanamycin, regenerated resistant plants were obtained. The NPT Ⅱ enzyme activity analysis, polymerase chain reaction (PCR), Southern blotting, Northern dot blotting and Western blotting demonstrated that the target gene was integrated into the genome of potato cells and well expressed in the plant.
