Objective To clone the full-length cDNA sequence of 9-cis-epoxycarotenoid dioxygenase (NCED) gene from leaves of Pyrus bretschneideri‘Xuehuali’ × P. pyrifolia ‘Shinsseiki’ cultivar ‘Huangguan’, study the relationship between its expression and the endogenous abscisic acid (ABA) content in leaves under different degree of drought, and provide a theoretical basis for understanding the molecular mechanism of the gene in drought resistance of pear tree.
Method According to the PbpNCED3 sequence obtained from the transcriptome data, the full-length cDNA sequence was cloned by homologous sequence amplification. The sequence was identified as belonging to the NCED family by bioinformatics analysis. Endogenous ABA content in pear leaves under different soil moisture conditions was determined by enzyme-linked immunosorbent assay (icELISA), and the expression of PbpNCED3 gene was measured by real time qPCR.
Result The PbpNCED3 cDNA sequence is 1 831 bp encoding 603 amino acids. The protein has four conserved histidines required for activity, two conservative domain (MIAHPKxDP and HDFAITE) characteristic of the NCED family and a RPE65 domain. There is a putative chloroplast-targeting peptide at N-terminal. The protein structure prediction shows that the protein sequence has 70% similarity to vp14 (NCED family protein) and it has a amphipathic α helix structure. PbpNCED3 gene expression is positively related with ABA content in pear leaves with a correlation coefficient of 0.914, and both of them are induced by drought.
Conclusion The PbpNCED3 gene obtained from the experiment belongs to the NCEDfamily. It responses to drought stress and its expression shares a similar dynamic trend with ABA content in pear leaves.
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