Age-related changes in relative expression of real-time PCR housekeeping genes in human skeletal muscle.
The purpose of this investigation was to examine the expression of three commonly used housekeeping genes -- glyceraldehyde-3-phosphate dehydrogenase (GAPDH), beta(2)-microglobulin (beta(2)M), and RNA polymerase 2a (polR2a) -- in elderly (E) compared to young (Y) subjects. Nine young subjects (22.7 +/- 3.4 yrs) and 11 elderly subjects (73.0 +/- 9.5 yrs) underwent a percutaneous skeletal muscle biopsy of the vastus lateralis. Equal concentrations of isolated mRNA from these samples were used to perform real-time polymerase chain reaction with primer/probe combinations specific to each gene of interest. The expression of GAPDH, beta(2)M, and polR2a was obtained as the value of cycle threshold (C(T)). An independent t-test with a level of significance at p < or = 0.05 was used to determine differences between groups. There was no difference in average C(T) of GAPDH between groups (p=0.869) (Y = 16.92 +/- 2.25 vs. E = 17.08 +/- 2.09) and polR2a (p = 0.089) (Y = 28.00 +/- 0.89 vs. E = 26.73 +/- 1.91). However, there was a significant difference (p < or = 0.05) in the average C(T) of beta(2)M (Y =21.79 +/- 0.44 vs. E = 21.05 +/- 0.51). The results indicate that special consideration needs to be made when selecting housekeeping genes for comparisons in real-time reverse-transcriptase polymerase chain reaction, depending upon the age of the populations of interest.
Touchberry
Chad D
CD
Applied Physiology Laboratory, University of Kansas, Department of Health Sport and Exercise Sciences, Lawrence, KS, USA.
Wacker
Michael J
MJ
Richmond
Scott R
SR
Whitman
Samantha A
SA
Godard
Michael P
MP
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000375:Aging; D005786:Gene Expression Regulation; D023281:Genomics / Q000379:methods*; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating) / Q000096:biosynthesis; D006801:Humans; D018482:Muscle, Skeletal / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D012319:RNA Polymerase II / Q000096:biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D001613:beta 2-Microglobulin / Q000096:biosynthesis
D012333:RNA, Messenger; D001613:beta 2-Microglobulin; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating); D012319:RNA Polymerase II