Biotechnology Bulletin ›› 2015, Vol. 31 ›› Issue (5): 120-127.doi: 10.13560/j.cnki.biotech.bull.1985.2015.05.019

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Cloning and Expression Analysis of Actin Gene in Cinnamomum camphora

Li Yongpeng, Zhang Liwei, Yao Yao, Huang Rui, Du Li   

  1. (School of Life Science and Technology,Nangyang Normal University,Nanyang 473000)
  • Received:2014-10-15 Online:2015-05-18 Published:2015-05-18

Abstract: As a house-keeping gene, Actin has been always being used as an internal standard to normalize mRNA levels between different samples by quantitative real-time PCR, thus plays an important role in gene expression analysis. In this research, through a method of homology cloning, a pair of degenerate primers were designed based on the conserved sequences of Actin genes from other plants submitted to GenBank, and then 4 cDNA fragments from Cinnamomum camphora were obtained using RT-PCR. Molecular biological analysis showed that, each of the 4 cDNA fragments was 998 bp and encoded a putative protein of 332 amino acids. Moreover, according to the homology analysis, the 4 cDNA fragments belonged to Actin subfamily, and they were named CcACTa, CcACTb, CcACTc and CcACTd, and deposited in GenBank(Accession number:KM086736 KM086737 KM086738 and KM086739). Quantitative real-time PCR results revealed that the expression level of CcACTc in different organs such as root, stem, leaf and leaves under low temperature treaments was relatively stable. It could serve as a candidate reference gene.

Key words: Cinnamomum camphora, Actin, gene cloing, expression analysis, quantitative real-time PCR