Molecular cloning and prokaryotic expression of a type II ribosome inactivating protein from Polyporus umbellatus
10.16438/j.0513-4870.2016-0731
- VernacularTitle:猪苓Ⅱ型核糖体失活蛋白A链基因的克隆以及原核表达
- Author:
Meng-meng LIU
1
;
Yong-mei XING
1
;
Shun-xin GUO
1
Author Information
1. Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100193, China
- Publication Type:ORIGINAL ARTICLES
- Keywords:
Polyporus umbellatus;
type Ⅱ ribosome inactivating protein;
gene clone;
expression analysis;
protein expression and purification
- From:
Acta Pharmaceutica Sinica
2017;52(2):327-332
- CountryChina
- Language:Chinese
-
Abstract:
A type Ⅱ ribosome inactivating protein (RIP) gene was cloned from Polyporus umbellatus sclerotia by RT-PCR method. The full open reading frame cDNA sequence of this gene was 873 bp in length and encoded a 290-aa protein with a molecular weight of 32.33 kDa and an isoelectric point of 5.58. Multiple sequence alignment revealed that the deduced amino acids possessed conserved domains of RICIN superfamily protein. A neighbor joining phylogenetic analysis suggests that PuRIP was closely related to RIP in Marasmius oreades. Real time PCR results showed that this gene expressed in all tested tissues of P. umbellatus. Meanwhile, the expression of this gene was significantly up-regulated in the part infected by Armillaria mellea. This result suggested that this PuRIP might played important role with potential biotic stress tolerance of P. umbellatus. Otherwise, we successfully constructed the pET15b-PuRIP plasmid, produced and purified the His-PuRIP fusion protein, which would provide the basic material for polyclonal antibody preparation and gene function research.
