Relative mRNA expression of the lactate dehydrogenase A and B subunits as determined by simultaneous amplification and single strand conformation polymorphism. Relation with subunit enzyme activity.
To explore if it is correlated in human tumor cells that the expression of LDH homologous gene and LDH isoenzymes, we used RT-PCR-SSCP technique to measure the relative expression of genes with homologous sequences. The combination of PCR using common primers designed in the highly conserved regions and single-strand conformation polymorphism analysis of the products is used for quantitative determination of the proportions of LDH-A mRNA in human cancer cell lines. The proportion is compared with that of the activities of isoenzymes. The results indicated that the enzyme activity of LDH-A was consistent with mRNA levels in the human tumor cell. The present procedure using a single pair of primers for two fragments can overcome disadvantages in quantitative analysis using multiplex PCR. Template concentrations and PCR cycles did not affect the proportions of LDH-A and LDH-B in the product.
Liu
Ze-Jun
ZJ
Department of Laboratory Medicine, Southwest Hospital, Third Military Medical University, 400038, Chongqing, PR China. a65424208@online.cq.cn
Peng
Wei-Cong
WC
Yang
Xin
X
Huang
Jun-Fu
JF
Zhang
Xiao-Bing
XB
Zhang
Yun
Y
Maekawa
Masato
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D045744:Cell Line, Tumor; D017931:DNA Primers; D018076:DNA, Complementary; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D007770:L-Lactate Dehydrogenase / Q000235:genetics*; D000080864:Lactate Dehydrogenase 5; D018807:Polymorphism, Single-Stranded Conformational; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D012333:RNA, Messenger; D007770:L-Lactate Dehydrogenase; D000080864:Lactate Dehydrogenase 5; C409667:lactate dehydrogenase 1