Selection of HPRT primers as controls for determination of mRNA expression in dogs by RT-PCR.
Reliable housekeeping gene controls are critical for measuring and comparing gene expression at the transcription level by Northern blot and RT-PCR. In order to develop such controls for studying cytokine mRNA expression in dogs, DNA sequence encoding a full-length canine HPRT protein has been obtained. Numerous primer pairs derived from the canine HPRT sequence have been tested on canine genomic DNA as well as cDNA. The data from the present study suggest that there may be processed HPRT pseudogenes in dogs. Three pairs of canine HPRT primers designed and tested in the present study were able to differentiate between cDNA and genomic DNA under specific PCR conditions. These primers would be useful controls for measurement of mRNA expression by RT-PCR in the dog.
Johnson
Christine M
CM
Heska Corporation, 1613 Prospect Parkway, Fort Collins, CO 80525, USA.
Yang
Shumin
S
Sellins
Karen S
KS
Frank
Glenn R
GR
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers / Q000737:chemistry / Q000235:genetics; D004285:Dogs / Q000235:genetics* / Q000378:metabolism; D007041:Hypoxanthine Phosphoribosyltransferase / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D011544:Pseudogenes; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D017931:DNA Primers; D012333:RNA, Messenger; D007041:Hypoxanthine Phosphoribosyltransferase