Cloning and Expression Analysis of the Pear Black Spot Resistant Gene PpEMS1

doi:10.13560/j.cnki.biotech.bull.1985.2019-0240

Abstract

Abstract: This study aims to clone the gene related to pear disease resistance and to preliminary analyze the effect of the gene on resistance and defense of pear black spot. Based on homologous gene cloning method,a gene predicted to be the receptor protein kinase EMS1 was cloned from the ‘Huang-guan’ pear,temporarily named as PpEMS1. The gene then was analyzed by bioinformatics,Southern Blot,subcellular localization and quantitative real-time PCR. As result,its CDS was 3 894 bp,containing an open reading frame of 3 891 bp and encoding 1 296 amino acids. Southern blot analysis indicated that this gene existed as a single copy in ‘Huang-guan’ pear,‘Huang-hua’ pear and ‘Cui-yu’ pear,while it existed as multiple copies in ‘Ai-dang’ pear. Subcellular localization demonstrated that the fusion protein was expressed in both cytomembrane and nucleus. The results from the quantitative real-time PCR revealed that the expression of the PpEMS1 gene increased first and then decreased in both ‘Huang-hua’ and ‘Ai-dang’ pears and was induced at 1 d,2 d,3 d,4 d,5 d,6 d,7 d and 8 d after inoculating black spot pathogen. It is inferred that the PpEMS1 gene may be associated with the pear black spot resistance.

Key words: pear, PpEMS1, southern blot, quantitative real-time PCR, subcellular localization

DUAN Min-jie, YI Hong-wei, WANG Jin, WU Zheng. Cloning and Expression Analysis of the Pear Black Spot Resistant Gene PpEMS1[J]. Biotechnology Bulletin, 2019, 35(11): 16-21.

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