安全环保的烟草质体多顺反子定点整合表达载体的构建

Construction of The Safely and Friendly Environmental Tobacco Chloroplast Multicistron Site Integration Expression Vector

根据已发表的序列,通过PCR技术克隆了一系列构建烟草叶绿体多顺反子表达载体所需的元件:质体核糖体(16S)RNA操纵元启动子(Prrn)、质体psbA基因3’端(psbA3’)、山菠菜甜菜碱醛脱氢酶基因(BADH)、烟草叶绿体高频同源重组片段(psaA/psbC,大小3463bp)、甘露聚糖酶基因(man)、绿荧光蛋白基因(gfp)。构建了烟草质体多顺反子定点整合表达载体pLM7(-psaA-Prrn-SD-man-SD-gfp-SD-BADH-psbA3’-psbC-)。并在大肠杆菌中通过平板定性分析等方法对所构建载体上的表达盒进行了功能鉴定。

According to the published DNA sequences, A serial of dements for construcfing the tobacco chloroplast mulficistron expression vectors have been cloned by PCR technique, including Prm （a modified plasfid ribosomal RNA operon promoter）, psbA3＇ （the 3＇ region of the plasfid psbA gene）, BADH gene （encoding hetaine aldehyde dehydrogenase, derived from mountain spinach）, man gene （encoding mannase）, gfp gene （encoding green flurescence protein） and tobacco chloroplast high- frequency homologous recombination ctDNA frag- ment （psaA/psbC, 3463bp） .A tobacco chloroplast multicistron expression vector pLM7 （ - psaA- Prrn - SD- man - SD- gfp - SD - BADH - psbA3＇ - psbC - ） was constructed with these elements.And the function of expression cassette was identified.