[Cloning of a homologous gene of Magnaporthe grisea PMK1 type MAPK from Ustilaginoidea virens and functional identification by complement in Magnaporthe grisea corresponding mutant].
OBJECTIVE
Cloning of a Homologous Gene of PMK1 Type Mitogen-Activated Protein Kinase (MAPK) from the rice false smut fungus Ustilaginoidea virens.


METHODS
According to the conserved amino acid sequence of several filamentous fungus MAPKs, which were homologous to Magnaporthe grisea PMKI, degenerate PCR primers were designed to amplify the MAPK internal DNA fragment from Ustilaginoidea grisea. The complete UVMK1 DNA and cDNA sequences were obtained using Thermal Asymmetric Interlaced-PCR (TAIL-PCR) and RT-PCR methods. Functional Identification was done by using the M. grisea APMKI mutant stain nn78, including appressoria differentiation assay and barley infection test.


RESULTS
The total length of UVMKJ was 1435 bp. It contained 3 introns and encoded 355 amino acids. The induced amino acid sequence showed identical to Magnaporthe grisea PMKI, Fusarium oxysporum FMKJ, Fusarium solani FsMAPK, Colletotrichum lagenarium CMKI, Botrytis cinerea BMKI, Claviceps purpurea CMPKI. After transformation of the APMK1 mutant of M. grisea using a complement vector with the complete cDNA of UVMK1 (under the M. grisea MPG1 promoter), five transformants were obtained. Furthermore, the selected two transformants fully restored their ability to form appressoria and infect a barley leaf.


CONCLUSIONS
In this study, we characterized the frst MAPK protein from U. virens, and that UVMK1 is a homologue of M. grisea PMK1.
Zhang
Zhen
Z
Institute of Plant Protection and Microbiology, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China. zzhangcn@gmail.com
Du
Xinfa
X
Chai
Rongyao
R
Wang
Jiaoyu
J
Qiu
Haiping
H
Mao
Xueqin
X
Sun
Guochang
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D002967:Claviceps / Q000201:enzymology; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000235:genetics* / Q000502:physiology; D020082:Magnaporthe / Q000145:classification / Q000235:genetics*; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D055330:Nectria / Q000201:enzymology; D010802:Phylogeny; D010935:Plant Diseases / Q000382:microbiology; D017386:Sequence Homology, Amino Acid
D005656:Fungal Proteins; D020928:Mitogen-Activated Protein Kinases; C102726:Pmk1 protein, Magnaporthe grisea