Cloning and Expression Analysis of Cold Shock Protein (CSP) Gene in Pinus massoniana
LI Xue-Ying1,2, XU Ying-Ying1,2, ZHAO Yuan-Xiang1,2, WU Feng1,2,*
1 Institute for Forest Resources and Environment of Guizhou/Key Laboratory of Forest Cultivation in Plateau Mountain of Guizhou Province, Guizhou University, Guiyang 550025, China; 2 College of Forest, Guizhou University, Guiyang 550025, China
Abstract：Cold shock protein (CSP) is an evolutionarily conserved DNA/RNA binding protein that plays a key role in the establishment of cold tolerance and growth, as well as in the development of plants. In order to understand the family characteristics of CSP gene (PmCSP) in Pinus massoniana, this study used full-length transcriptome data to identify the PmCSP gene family and to analyse its phylogeny, structure and functions, as well as the expression patterns of seedlings in response to cold stress. The results showed that 2 PmCSP gene family members were identified and cloned, both were intronless. The open reading frame of PmCSP1 was 642 bp in length, encoding 213 amino acids residues. The open reading frame of PmCSP2 was 603 bp in length, encoding 200 amino acids residues. The encoded polypeptide chains contained both cold shock domains and CCHC zinc finger domains, but differed in the number of zinc finger domains. PmCSP1 contained 3 domains, while PmCSP2 only contained 2 domains. Phylogenetic analysis showed that PmCSPs were more closely related to P. tabuliformis. Protoplast transient expression analysis showed that PmCSPs proteins were mainly located in the nucleus and cytoplasm. The expression of PmCSPs in the leaves and terminal buds of P. massoniana seedlings were higher than those in other tissues. Under cold stress, PmCSP1 and PmCSP2 were upregulated in the primary needle and secondary needle seedlings, respectively, with PmCSP expression peaking at 8 h, which indicated that PmCSPs were early response genes for P. massoniana seedlings in response to cold stress and may be involved in specific molecular regulation in heteroblastic foliage seedlings. This study provides a theoretical basis for further revealing the functions of the PmCSP gene family and the regulatory mechanisms established for the cold resistance of conifer heteroblastic foliage seedlings.
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