[Construction of eukaryotic expression vector of rat HAS-3 gene and its effects on chemotaxis].
OBJECTIVE
To construct the eukaryotic expression vector of rat HAS-3 gene, express it in RSC96 cells and detect the enzyme activity of the recombinant protein.


METHODS
Rat HAS-3 gene was cloned from CCI rat injury nerve cDNA using RT-PCR and inserted into pcDNA3.1D and pEGFP-N1. The recombinant plasmids were transfected to RSC96 cells. The expression level and enzyme activities of the recombinant proteins were monitored.


RESULTS
Rat HAS-3 gene was cloned into pcDNA3.1D and pEGFP-N1 correctly. Recombinant proteins were detected in RSC96 cells and the synthesis of HA was up-regulated after transcfection. The condition medium of RSC96 cells overexpressing HAS-3 had some effects on chemotaxis of macrophages.


CONCLUSIONS
The eukaryotic expression vectors of rat HAS-3 has been constructed successfully. The transfection of HAS-3 gene to RSC96 cells is effective in chemotaxis of macrophages which demonstrates HAS-3 may contribute to the development of inflammation in vivo.
Nie
Yong
Y
Department of Biological Science and Biotechnology, Tsinghua University, Beijing 100084, China. nieyong@gmail.com
Cai
Guo-Ping
GP
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D002633:Chemotaxis / Q000187:drug effects*; D017077:Culture Media, Conditioned / Q000378:metabolism / Q000494:pharmacology; D005822:Genetic Vectors / Q000235:genetics*; D014453:Glucuronosyltransferase / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000076002:Hyaluronan Synthases; D008264:Macrophages / Q000166:cytology / Q000187:drug effects; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*
D017077:Culture Media, Conditioned; D011993:Recombinant Fusion Proteins; D014453:Glucuronosyltransferase; C491355:Has3 protein, rat; D000076002:Hyaluronan Synthases