[Construction and expression of recombinant adenovirus carrying human SHP-1 gene].
OBJECTIVE
To construct and identify recombinant adenovirus Ad-SHP1 carrying human SH2 domain-containing protein tyrosine phosphatase (SHP-1).


METHODS
Human SHP-1 cDNA from healthy volunteers' peripheral blood lymphocyte was cloned into the shuttle plasmid pAdTrack-CMV by standard procedure. The plasmid pAdTrackCMV-SHP1 was selected and then linearized by Pme I, followed by homologous recombination with bone plasmid pAdEasy-1 in BJ5183. Recombinant adenovirus Ad-SHP1 were obtained after packaged in HEK293 cells and the green fluorescence in HEK293 cells was observed. Recombinant adenovirus was confirmed with PCR and the expressed SHP-1 protein was verified with Western Blotting. Ad-SHP1 was multiplied and purified. The titer of virus was measured with fluorescent counter. Results The results of sequencing, restriction endounclease digestion and Western Blotting indicated that Ad-SHP1 was successfully constructed. The titer of Ad-SHP1 reached 1.58 x 10(10) pfu/mL.


CONCLUSIONS
Recombinant adenovirus Ad-SHP1 had been successfully constructed. It could express SHP-1 protein stably and effectively and this will be very helpful for the further study of the generation of extranodal NK/T cell lymphoma and the corresponding therapy.
Wang
Xiao-ling
XL
Department of Pathology, West China Hospital, Sichuan University, Chengdu 610041, China.
Zhao
Sha
S
Yu
Jian-bo
JB
Huang
Rui
R
Zhang
Ying-chun
YC
Liu
Wei-ping
WP
D000256:Adenoviridae / Q000235:genetics* / Q000502:physiology; D015153:Blotting, Western; D045744:Cell Line, Tumor; D004274:DNA, Recombinant / Q000235:genetics*; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D006801:Humans; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6 / Q000235:genetics*; D017422:Sequence Analysis, DNA; D019562:Viral Load
D004274:DNA, Recombinant; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6