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水稻Rubisco小亚基前体cDNA的克隆及其产物向豌豆叶绿体的运输

陈根云, 颜日辉, 李立人

中科院上海植物生理研究所.上海 200032

摘要

利用RT-PCR方法克隆了含完整前导肽的水稻Rubisco小亚基前体cDNA,经体外转录和翻译合成同位素标记的prSS,然后在体外进行了对豌豆完整叶绿体的穿膜运输,并观察了光、ATP等对prSS穿膜运输的影响。

关键词： Rubisco小亚基; RT－PCR; 体外转录／翻译; 蛋白质运输; 叶绿体

Cloning of rice rbcS precursor cDNA and the import of its in vitro synthesised products into intact chloroplasts from pea

Chen Genyun, Yan Rihui, Li Liren

Shanghai Institute of Plant Physiology,Chinese Academy of Sciences.Shanghai 200032

Abstract

The precursor cDNA of Rubisco small subunit (rbcS) from rice was cloned in pSPT19 by RT-PCR. The results of sequencing indicated that the fragment of rbcS contained the full length transit peptide. By comparing its sequence of transit peptide with that of homology of the transit peptide sequence between the two species is only 39.5%, but they have three high conservation regions, in which the percentage of homologues is up to 80%. Furthermore, the rice precursor of rbcS (prSS) labeled with ~(35)S-Met was synthesised in vitro from plasmid-transcribed mRNA in wheat germ system (cell free). After being incubated with fresh intact chloroplasts from pea, rice prSS was shown to be able to cross the membrane of pea chloroplasts. The amount of the prSS imported into chlroplast under illumination was 17% more than that treated in the dark; and the amount was increased by 80% when the chlorplasts is treated with ATP, ADP had no obvious effect on the import. Furthermore, the amount of SS imported into the chloroplasts was directly proportional to the amount of prSS added in the import assays.

Key words: PrSS;RT-PCR;Synthesised in vitro;Protein import;Chloroplasts

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引用本文：

陈根云, 颜日辉, 李立人. 水稻Rubisco小亚基前体cDNA的克隆及其产物向豌豆叶绿体的运输[J]. 生理学报 1998; 50 (3): .

Chen Genyun, Yan Rihui, Li Liren. Cloning of rice rbcS precursor cDNA and the import of its in vitro synthesised products into intact chloroplasts from pea. Acta Physiol Sin 1998; 50 (3): (in Chinese with English abstract).