| Title | [cDNA cloning and location of Pagumogonimus skrj abini cysteine protease] | | Author(s) | Wang Y, Zhang XL, Zhang YL, Duan JH, Zhang JR, Huang FS | | Institution | Department of Pathogenic Biology, Third Military Medical University, Chongqing 400038. | | Source | Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi 2003; 21(4):214-7. | | MeSH | Amino Acid Sequence
Animals
Base Sequence
Cloning, Molecular
Cysteine Endopeptidases
DNA, Complementary
English Abstract
Gene Expression
In Situ Hybridization
Molecular Sequence Data
Paragonimus
Reverse Transcriptase Polymerase Chain Reaction
| | Abstract | OBJECTIVE: To clone the cysteine protease cDNA fragment from Pagumogonimus skrjabini adults and locate the tissue of the adult worm where cysteine protease is expressed.
METHODS: The cysteine protease cDNA fragment was amplified by reverse transcription-polymerase chain reaction (RT-PCR) with degenerated primers. The production was TA-cloned into the pUCm-T vector and sequenced. DNASIS program was used to analyse the nucleotide sequence and deduce the amino acid sequence, which was aligned with the correlated parasite cysteine protease afterwards. The digoxin labeled cRNA probe was synthesised by in vitro transcription with the cloned cDNA as template. The frozen sections of the adult worms were analysed by hybridization in situ to locate the gene expression.
RESULTS: A 495 bp cDNA fragment was amplified by RT-PCR and sequenced. An amino acid sequence was deduced by DNASIS. Sequence analysis and alignment showed significant homologies with the correlated parasite cysteine proteinases and conservation of Cys, His and Asn residues that from a catalytic triad. In the hybridization in situ analysis, intestinal epithelium was stained positively on transverse section of adult worms.
CONCLUSION: The cysteine proteinase cDNA fragment from Pagumogonimus skrjabini adults was cloned. There are some key sites which are correlated to the function of cysteine protease in the cDNA fragment. Cysteine protease is mainly expressed in intestinal epithelium of P. skrjabini. | | Language | chi | | Pub Type(s) | Journal Article
| | PubMed ID | 14628371 |
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