墨兰‘达摩’原叶绿素酸酯氧化还原酶基因CsPORB功能的初步研究

Preliminary Study on the Function of NADPH: Protochlorophyllide Oxidoreductase (CsPORB) Gene in Cymbidium sinense ‘Dharma’

摘要:

为了解墨兰‘达摩’（Cymbidium sinense ‘Dharma’）的POR基因功能，从其嫩叶中克隆得到CsPORB基因。CsPORB的开放阅读框长度为1 200 bp，编码399个氨基酸，CsPORB分子量为43.13 kDa，理论等电点9.30；CsPORB与其他物种的POR蛋白序列高度同源，在进化关系上较为保守；qRT-PCR表达分析表明，CsPORB在墨兰不同发育阶段和不同组织中的表达有明显差异，其集中于生长旺盛的嫩叶中表达。经高表达转基因表型验证，35S：CsPORB转拟南芥（Arabidopsis thaliana）植株中PORB及其下游叶绿素合成代谢酶活性显著提高，叶绿素含量增加，尤其在黑暗处理1周后，35S：CsPORB转基因植株仍表现持绿表型。因此，CsPORB基因可能在墨兰‘达摩’叶绿素合成代谢中发挥重要作用。

Abstract:

In order to understand the function of POR gene in Cymbidium sinense ‘Dharma’, one POR gene family member CsPORB was cloned from young leaves. The CsPORB gene contained an open reading frame of 1 200 bp, which encoded a protein of 399 amino acids with molecular weight of 43.13 kDa and theoretical isoelectric point of 9.30. Sequence alignment showed that CsPORB was highly homologous to the sequence of POR family protein of other species, indicating conservative evolution of this gene family. The CsPORB expression showed obviously difference in different developmental stages and tissues, and high expression in growing young leaves. Furthermore, both of the PORB and its downstream chlorophyll biosynthesis enzyme activity increased in transgenic 35S:CsPORB Arabidopsis, as well as chlorophyll content, especially 35S:CsPORB transgenic plants remained green under dark for one week. Therefore, CsPORB could play an important roles in chlorophyll synthesis pathway of Cymbidium sinense ‘Dharma’.