[Construction of eukaryotic expression vector of unknown KH gene and its effect on cell proliferation].
OBJECTIVE
To construct an eukaryotic expression vector of open reading frame of unknown KH gene (KH-ORF), and investigate its effect on cell proliferation.


METHODS
The pCI-neo-KH-ORF expression vector was constructed by DNA recombinant technique and was introduced into COS-7 cells and K562 cells by lipofectactin-mediated DNA transfection. Expression of KH-ORF mRNA was detected by RT-PCR. The effect of KH-ORF on cell cycle of COS-7 cells and K562 cells was evaluated by flow cytometry (FCM). Effect on cell proliferation of COS-7 cells was tested by MTT assay and that on K562 cells was analyzed by growth curves and LDH activity measurement.


RESULTS
(1) KH-ORF mRNA was expressed both in COS-7 cells and K562 cells. (2) The cell cycle and cell proliferation of COS-7 cells were unaffected significantly. (3) The proportion of cells in S phase was increased in pCI-neo-KH-ORF-transfected K562 cells; and growth curves and LDH activity indicated enhanced cell proliferation.


CONCLUSIONS
KH gene may be a leukemia gene related to proliferation of K562 cells.
He
Yu-juan
YJ
Department of Clinical Biochemistry, Chongqing University of Medical Sciences, Chongqing 400016, China.
Ou
Yi-heng
YH
Zhang
Yan
Y
Liu
Xiao-shan
XS
Ma
Ling-di
LD
Jiang
Ji-kai
JK
D000818:Animals; D019556:COS Cells; D049109:Cell Proliferation; D002522:Chlorocebus aethiops; D015972:Gene Expression Regulation, Neoplastic; D052138:Genes, Neoplasm / Q000235:genetics* / Q000502:physiology; D005822:Genetic Vectors; D006801:Humans; D020014:K562 Cells; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D016366:Open Reading Frames / Q000235:genetics* / Q000502:physiology; D010957:Plasmids; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics; D016196:S Phase; D014162:Transfection
D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D007770:L-Lactate Dehydrogenase