黄冠梨几丁质酶基因家族cDNA全长序列克隆与分析

根据已知植物几丁质酶基因序列的保守区域，设计特异性引物，以梨黄冠品种叶片为材料，提取RNA，进行反转录，克隆获得5个几丁质酶基因cDNA片段，进一步通过RACE技术获得该基因的全长cDNA序列，依次命名为PyChi1、 PyChi2、 PyChi3、 PyChi4、 PyChi5，序列提交GenBanK，登录号为：FJ589783、 FJ589784、 FJ589785 、 FJ589786、FJ589787。序列分析结果表明，所获得5个几丁质酶基因，都具有完整的ORF和Poly(A)结构，与其他植物几丁质酶基因具有较高的相似性；5个基因的核酸序列的同源性约在35%~53%之间，PyChi1和PyChi4的氨基酸同源性最高，约85%，而与PyChi5约77%；在几丁质酶基因家族，分别属于Chitinase ClassesⅠ-Ⅴ，为5个不同类型的成员；不同类型的几丁质酶基因在核酸和氨基酸序列以及功能结构方面都存在着存在着显著的差异，主要表现在肽链特性和基因结构等方面。这为进一步研究梨几丁质酶的结构和功能奠定了基础。

Full-length cDNA Clone and Sequence Analysis of the Chitinase Family Gene from Pear

In order to get the chitinase gene from pear,specific primers were designed based on the conserved sequence of the plant chitinase gene.Five different cDNA fragments of chitinase genes were amplified by RT-PCR from pear(Pyrus hybrid cv.Huangguan).The full-length cDNAs of Huangguan chitinase genes were produced by using RACE,and named PyChi1,PyChi2,PyChi3,pyChi4 and PyChi5(GenBank accession No.:FJ589783,FJ589784,FJ589785,FJ589786 and FJ589787),respectively.The results showed that the PyChi1～5 were all full-length cDNA sequence including the complete open reading frame(ORF)and the structure Poly(A),and had higher identities with other plant chitinase gene.Among the five chitnase genes,the nucleic acid sequence identity was 35%～53%,PyChi1 and PyChi4 had the highest amino acid sequence identity,which was 85%,while PyChi1 and PyChi5 produced the lowest amino acid sequence identity.which was 49%.The five chitinaes belong to chitinase Classes Ⅰ～Ⅴ in the classification of cllitinase family,respectively.Using the bioinformatics analysis,the five types of chitinase had obviously different in sequence of nucleic acid and amino acid,especially about the characteristics of peptide and chitnnase gene structure.These results are helpful for further studies on structure and function of pear chitinase.