Molecular Cloning and Characterization of TGF-β and Its Response to Seawater Acidification in the Sea Urchin Meconocrotus nudus

doi:10.13560/j.cnki.biotech.bull.1985.2018-0470

Abstract

Abstract:

This work aims to clarify the sequences and structure of transforming growth factor-β gene（TGF-β）and explore its expression mode under seawater acidification. We used homology-based cloning and rapid amplification of cDNA ends（RACE）to acquire the full-length cDNA sequence of novel gene TGF-β from Meconocrotus nudus（here designated MnTGF-β）. Results are as such：（1）The full-length cDNA of MnTGF-β were 2 299 bp,the length of 5'noncoding region was 745 bp,the length of 3'noncoding region was 261 bp,the length of ORF was 1 290 bp,and it encoded 430 amino acid with the relative molecular weight of 48.31 kD,and the theoretical pI of 5.34. （2）Homology and phylogenetic analysis showed that MnTGF-β protein sequence was highly conserved with SpTGF-β2 protein sequence in Strongylocentrotus purpuratus（homology was 97.69%）. （3）Quantitative real-time PCR analysis demonstrated that MnTGF-β was ubiquitously expressed in gonad,coelomic fluid,intestines,peristomial membrane and tube foot of healthy adult M. nudus,and the order of its expression was as tube foot > peristomial membrane > intestines > coelomic fluid >gonad.（4）Compared with the natural seawater group（pH 8.06 ±0. 01）,when △pH was -0. 3,the relative expression of gene MnTGF-β in the gonad of sea urchin tended to increase first,then decrease significantly（P < 0. 01）,and then increased significantly（P < 0. 01）with the increase of acidification time;the relative expression level in tube foot was significantly up-regulated（P < 0. 01）,but that in intestines significantly decreased（P < 0. 01）and then increased（P < 0. 05）. When the △pH in seawater were -0. 4 and -0. 5,the relative expression of gene MnTGF-β in the gonads and tube foot of sea urchins showed a tendency of first decreasing and then increasing with the prolongation of acidification time,while that in the intestines decreased significantly（P < 0.01）.

Key words: Meconocrotus nudus, TGF-&#x003b2, gene cloning, seawater acidification, expression mode

LI Ying-ying, CUI Dong-yao, CHANG Ya-qing, LIU Lin, ZHANG Bao-jing, JIANG Hui-ting, LIU Yin, ZHAN Yao-yao. Molecular Cloning and Characterization of TGF-β and Its Response to Seawater Acidification in the Sea Urchin Meconocrotus nudus[J]. Biotechnology Bulletin, 2019, 35(2): 116-128.

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