摘要
从担子菌灰树花的菌丝体中提取总 RNA,并纯化出 m RNA,m RNA经反转录合成c DNA第一链 ,以 c DNA第一链为模板经 PCR扩增海藻糖合酶 ( Tsase)基因 ,获得一长约2 .2 kb的片段 ,把该片段连接到 p GEM○R- T- easy vector上进行测序 ,其全长共 2 1 99bp.随后将此片段以正向插入植物表达载体 p BI1 2 1的 35S启动子和 NOS终止子之间 ,构建了一植物表达载体 p BT;又把 ubi- L启动子插入 p BI1 2 1的 H ind + Xbal位点构建 p UB,再把海藻糖合酶基因以正向插入载体 p UB的 Bam H1 + Sst1位点构建了另一植物表达载体 p UBT.
The total RNA from bacidiomycete, Grifola frndosa was isolated. The mRNA was purified and reverse transcripted to first strand of cDNA. A pair of primers were designed and synthesized according to the sequence reported. The very specific PCR product of trehalose synthase gene was obtained by using the first strand of cDNA as template. The product was ligased to pGEM ○[KG-2/3]R T easy vector and sequenced. The sequencing data showed that the PCR product was 2 199 bp. The Tsase gene sequence was then inserted between the CaMv 35S promoter and NOS terminator into the expression vector pBI121. This expression vector was called pBT; The ubi L promoter was first inserted into the sites of the expression vector pBI121/HindIII+Xbal, and this recombinant plasmid was called pUB, then the Tsase gene was inserted into the sites of the plasmid pUB/BamHI+SstI. Hence the expression vector pUBT was constructed.
出处
《生命科学研究》
CAS
CSCD
2000年第2期130-135,共6页
Life Science Research
基金
国家自然科学基金资助项目!(397690 0 1 4 )
