Construction of a subtracted cDNA library of chronic intermittent hypoxia rabbit liver by suppression subtractive hybridization.
- Author:
Yue-tao WU
1
;
Rui-hong LIU
;
Yu YANG
;
Ying-quan LUO
;
Yao RONG
Author Information
1. Department of Geriatrics, Second Xiangya Hospital, Central South University, Changsha 410011, China. wuyutao99@126.com
- Publication Type:Journal Article
- MeSH:
Animals;
Gene Library;
Genetic Vectors;
Hyperlipidemias;
Hypoxia;
metabolism;
pathology;
Liver;
metabolism;
pathology;
Nucleic Acid Hybridization;
methods;
RNA, Messenger;
genetics;
Rabbits
- From:
Journal of Central South University(Medical Sciences)
2007;32(6):1013-1019
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To construct a subtracted cDNA library of chronic intermittent hypoxia (CIH) rabbit liver by suppression subtractive hybridization (SSH).
METHODS:Twenty-four rabbits were divided into 4 groups: ordinary feeding group, full-fat food group, ordinary feeding in chronic intermittent hypoxia group, and full-fat food in chronic intermittent hypoxia group. The mRNAs were extracted from different rabbit livers and converted into double-strand cDNA. After digestion with restriction enzyme, the cDNA of hyperlipidemia-sensitive rabbit group was subdivided into 2 portions and each one was lighted with different adaptors. Two rounds of both hybridization and suppression PCR obtained the differentially expressed cDNA. The PCR products were inserted into T/A vector to set up the subtractive cDNA library. The clones were selected and amplified by PCR and identified.
RESULTS:Based on the pathology of the abdominal aorta and liver, and the amplified library contained 500 positive bacteria clones, including 462 clones, which had inserts from 250 to 700 bp by PCR analysis. A novel rabbit gene, Cthrc1, involved in CHI had been cloned. The GenBank Accession Number is XM_418373.
CONCLUSION:The molecular mechanism of CIH promoting atherogenesis formation is made clear.
