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Canine glyceraldehyde-3-phosphate dehydrogenase complementary DNA: polymerase chain reaction amplification, cloning,
A Gröne1, M T Weckmann, C C Capen
1Department of Veterinary Biosciences, College of Veterinary Medicine, Ohio State University, Columbus 43210, USA.
Objective:
To use canine glyceraldehyde-3-phosphate dehydrogenase complementary DNA (GAPDH cDNA) as a template in ribonuclease (RNase) protection assays to measure canine GAPDH mRNA expression.
Design And Procedure:
Primers designed from the human GAPDH gene were used to amplify a 191-base pair canine GAPDH cDNA by reverse-transcription polymerase chain reaction. The cDNA was sequenced, and used as a template for RNase protection assay.
Sample Population:
Total RNA was isolated from a canine squamous carcinoma cell line.
Results:
Canine GAPDH cDNA had a high degree of homology to human, rat, and mouse GAPDH. In vitro transcription of canine GAPDH cDNA was used to produce complementary RNA that detected canine GAPDH mRNA by RNase protection assay.
Conclusion:
Canine GAPDH cDNA is a useful loading control to be used in RNase protection assays measuring mRNA expression in canine cells or tissues.