[Cloning and characterization of a homologous gene of plant class V chitinase from balsampear, Momordica charantia Linn].
Balsampear (M. charantia Linn.) is a vegetable crop, highly resistant to pathogens. Chitinases were proposed to play an important role in the defense response of this crop. Based on the N-terminal sequence of a purified balsampear chitinase, a fragment (ChitB), similar to the tobacco class V chitinase gene, was amplified from the leaf RNA using 3'RACE, and the corresponding 5' sequence was further amplified by the Y-RACE method. By joining the two amplified fragments, the full-length cDNA of M. charatica homologous gene of plant class V chitinase (McChi5) was obtained. The 1348 bp cDNA contained an ORF of 1044 bp, which coded for a polypeptide of 347 amino acids. The deduced polypeptide had a predicted molecular weight of 38.3 kD and a pI of 5.77. Homology analysis demonstrated that, McChi5 protein, which contained a conserved domain of family 18 glycosyl hydrolyse, had the sequence similar to tobacco class V chitinases, several putative chitinases and chitinase-like proteins of Arabidopsis thiliania, and some chitinases from mammals, insects and bacteria. Southern blotting suggested that two copies of McChi5 gene and several homologous genes existed in the M. charatica genome. By RNA dot blotting analysis, expression of the McChi5 gene was detected in cotyledons, roots, stems, and leaves, and it was not induced by wounding treatment. The biological functions and the potential applications of Mochi5 gene were discussed.
Xiao
Yue-Hua
YH
Biotechnology Research Center, Southwest Agricultural University, Chongqing 400716, China.
Hou
Lei
L
Yuan
Xiao-Hong
XH
Yang
Xing-Yong
XY
Pei
Ying
Y
Luo
Xiao-Ying
XY
Pei
Yan
Y
D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D002688:Chitinases / Q000235:genetics*; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics / Q000378:metabolism; D018628:Gene Dosage; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D029777:Momordica charantia / Q000201:enzymology / Q000235:genetics*; D010802:Phylogeny; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA
D018076:DNA, Complementary; D018744:DNA, Plant; D018749:RNA, Plant; D004262:DNA Restriction Enzymes; D002688:Chitinases