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转运肽对于大多数蛋白转运到叶绿体是必需的。虽然利用生物信息学分析可预测蛋白的定位信息及转运肽的序列信息,但转运肽的转运效果仍需要进一步的验证。本研究基于Gen Bank所报道的番茄Rubisco小亚基叶绿体转运肽(TCTP)的信息,利用特异引物从番茄DNA中扩增获得一段约170bp的片段,克隆到pMD~@18-T simple载体,测序表明获得番茄Rubisco小亚基的转运肽。为了进一步验证其功能,将其连接到瞬时表达载体P322-d1-eGFP,构建瞬时表达载体TCTP-eGFP-d1,利用PEG介导法将重组瞬时表达载体转入水稻原生质体,激光共聚焦显微镜分析表明,该转运肽可以顺利将eGFP定位到叶绿体,该研究有助于TCTP的进一步应用。
Abstract：Transit peptides are necessary for most proteins to be transported to the chloroplast. The targeting information of proteins and sequence information of transit peptides could be predicted by bioinformatics analysis,but the translocation effect of transit peptides still needs further verification. This research was based on the information of the Rubisco small subunit chloroplast transporter peptide(TCTP) reported by Gen Bank. A fragment about170 bp was amplified from tomato(Solanum lycopersicum) genomic DNA by using the specific primers, and cloned into p MD@18-T simple vector. The sequencing result showed that the transit peptide was obtained from the Rubisco small subunit of tomato. To further verify the functions of TCTP, it was connected to the transient expression vector P322-d1-e GFP, and the transient expression vector TCTP-e GFP-d1 was constructed. The recombinant transient expression vector was transferred into the protoplast of rice by PEG-mediated method. The laser confocal microscope analysis showed that the TCTPcould successfully locate e GFP into the chloroplasts, which could be helpful to the further application of TCTP.
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基本信息:
DOI：10.13417/j.gab.037.001570
中图分类号:Q943.2
引用信息:
[1]胥华伟,侯典云.番茄Rubisco小亚基叶绿体转运肽的克隆及其功能验证[J].基因组学与应用生物学,2018,37(04):1570-1575.DOI:10.13417/j.gab.037.001570.
基金信息:
国家自然科学基金(31100197);; 河南科技大学博士启动基金(13480027)共同资助
2018-04-25
2018-04-25