从东方山羊豆（Galega.orientalisL.）中克隆出的GoMIPS（肌醇-1-磷酸合成酶）基因序列设计合成1对带有NcoI和SpeI酶切位点的引物，用RT-PCR方法从东方山羊豆幼嫩叶片总RNA中扩增出GoMIPS基因的cDNA序列，经纯化后，克隆至pMD19-T载体上，构建pMD-MIPS的中间载体，测序鉴定。然后用NcoI和SpeI限制性酶对含有目的基因的pMD-MIPS和pCAMBIA1302空载体进行双酶切，通过酶切鉴定和测序分析表明，已成功构建了CaMV35S启动子驱动GFP报告基因的双元植物表达载体pCAMBIA1302-MIPS。
Abstract
A pair of primers containing the restriction sites for NcoI/SpeI were designed according to the cloned GoMIPS (inositol-1 - phosphate synthase) gene sequence of Galega orientalis. The GoMIPS cDNA was amplified by RT-PCR from the total RNA of the young leaves of the Oriental Galega. After purification, RT-PCR products were cloned to the pMD19-T vector were dealt to construct the pMD-MIPS in the middle of the carrier DNA sequencing. pMD-MIPS containing the target gene and pCAMBIA1302 empty vector with NcoI / SpeI restriction enzyme digestion,the restriction endonuclease and sequence analysis shows that a dual CaMV35S promoter-driven GFP reporter geneplant expression vector pCAMBIA1302-MIPS has been successfully constructed.
关键词
东方山羊豆（Galega.orientalisL.） /
GoMIPS /
pMD19-T /
pCAMBIA1302
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Key words
Galega. orientalis L. /
GoMIPS (inositol -1- phosphate synthase) /
pMD19-T vector /
pCAMBIA1302 vector
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参考文献
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脚注
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基金
山西省农业科学院畜牧兽医研究所基金项（YGG0853）
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