The Gene Cloning and Expression Analysis of SCPL in Tea Plant (Camellia sinensis)

QIU Chuan-hui; LI Wei-wei; WANG Yun-sheng; LI Ming-zhuo; LUO Yang; LIU Ya-jun; GAO Li-ping; XIA Tao

doi:10.13305/j.cnki.jts.2013.03.011

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Journal of Tea Science ›› 2013, Vol. 33 ›› Issue (3) : 202-211. DOI: 10.13305/j.cnki.jts.2013.03.011

The Gene Cloning and Expression Analysis of SCPL in Tea Plant (Camellia sinensis)

QIU Chuan-hui¹, LI Wei-wei², WANG Yun-sheng², LI Ming-zhuo¹, LUO Yang², LIU Ya-jun², GAO Li-ping^2,*, XIA Tao^1,*

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Abstract

In recent years, serine carboxypeptidase-like proteins (SCPL) have been found that they are involved in plant secondary metabolites with the function of transferring acyl. The full-length cDNA of three CsSCPL were cloned from Camellia sinensis by RT-PCR technology. Bioinformatics analysis showed that the three CsSCPL proteins contained SCPL family's characteristic structures, such as one substrate binding and three catalysis conserved regions, a number of N-glycosylation sites and a conserved catalytic triad Ser-Asp-His amino acid active catalytic site and so on. The phylogeny analysis showed that CsSCPL probably possessed acyltransferase function. Quantitative RT-PCR analysis showed that the CsSCPL genes expressed in bud, leaf, stem and root. The relative expression of CsSCPL1 and CsSCPL3 in the leaves was significantly higher than that in stems and roots, while CsSCPL2 was highly expressed in the root. The CsSCPL genes were constructed into expression vector pET-32a(+) for over expression in prokaryotic cells and optimal inducing conditions including time, temperature were studied. The SDS-PAGE showed that recombinant proteins with formula weight 70βkD were induced successfully by IPTG, which coincided with the prediction.

Key words

Camellia sinensis / bioinformatics analysis / prokaryotic expression / serine carboxypeptidase-like proteins

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QIU Chuan-hui, LI Wei-wei, WANG Yun-sheng, LI Ming-zhuo, LUO Yang, LIU Ya-jun, GAO Li-ping, XIA Tao. The Gene Cloning and Expression Analysis of SCPL in Tea Plant (Camellia sinensis)[J]. Journal of Tea Science. 2013, 33(3): 202-211 https://doi.org/10.13305/j.cnki.jts.2013.03.011

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