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通过对刚毛柽柳转录组的分析,获得并克隆了1个H~+-PPase基因cDNA全长,命名为ThVP3。序列分析结果表明:ThVP3全长3 500 bp,包含2 406 bp完整的开放读码框,618的5’非编码区和476 bp的3’非编码区。该cDNA编码1个801个氨基酸的多肽,等电点为5.67,推测分子质量为84.86 k D。通过与其他物种的氨基酸多序列比对及系统进化树分析,发现ThVP3氨基酸序列与AtVP2(拟南芥)一致性达88%,属于Ⅱ型VP成员。ThVP3基因有15个跨膜区,并具有高度保守的3个结构域(CS1,CS2,CS3)。相对荧光定量RT-PCR结果表明,ThVP3在高盐及干旱胁迫下均呈现为表达上调,但在各胁迫时间点和不同组织中的表达模式有所差异,0.4 mol/L Na Cl胁迫处理下,地下部分3 h达到最大值后有不同程度的降低,地上部分24 h达到最大值,与地下部分比,地上部分呈现较高的表达量;20%(w/v)PEG 6000胁迫处理下,地下部分在3 h和12 h达到峰值,地上部分表达量逐渐升高,24 h达到最大值后逐渐降低,96 h达到第2个峰值。ThVP3的表达受盐旱胁迫的诱导,推测其可能在柽柳的胁迫应答中起着重要的作用。
Through analyzing the transcriptome of Tamarix ramosissima, one full-length cDNA of H ~ + -PPase gene was obtained and cloned, named ThVP3. Sequence analysis showed that ThVP3 was 3 500 bp in length and contained 2 406 bp complete open reading frame, 618 ’5’ non-coding region and 476 bp 3 ’non-coding region. This cDNA encodes a polypeptide of 801 amino acids with an isoelectric point of 5.67 and a predicted molecular mass of 84.86 kD. Through the amino acid sequence comparison and phylogenetic tree analysis with other species, the amino acid sequence of ThVP3 was found to be 88% identical to that of AtVP2 (Arabidopsis thaliana) and belongs to the type II VP. The ThVP3 gene has 15 transmembrane domains and has three highly conserved domains (CS1, CS2, CS3). Relative fluorescence quantitative RT-PCR results showed that ThVP3 was up-regulated under salt stress and drought stress, but different expression patterns at different stress time points and different tissues were different. Under 0.4 mol / L NaCl stress, The underground part reached the maximum at 3 h and decreased to a certain extent, the aboveground part reached the maximum at 24 h, and showed a higher expression level than the underground part under the stress of 20% (w / v) PEG 6000. Some of them reached the peak at 3 h and 12 h, the expression of aboveground part increased gradually, reached the maximum at 24 h and then decreased gradually, reaching the second peak at 96 h. The expression of ThVP3 was induced by salt and drought stress, suggesting that it may play an important role in the stress response of Tamarix.