A sensitive, quantitative assay for measurement of allele-specific transcripts differing by a single nucleotide.

We have found that the single nucleotide primer extension assay, a PCR-based assay currently used qualitatively to measure allelic differences in DNA, can be used quantitatively to measure allele-specific transcripts differing by only a single nucleotide. We show that total RNA containing the Pgk-1a transcript can be specifically detected even in a 1000-fold excess of RNA containing the Pgk-1b transcript.

[1]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[2]  S. Spitzer,et al.  Single nucleotide primer extension to detect genetic diseases: experimental application to hemophilia B (factor IX) and cystic fibrosis genes. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[3]  J. Sklar,et al.  Rapid, nonradioactive detection of clonal T-cell receptor gene rearrangements in lymphoid neoplasms. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[4]  Robert L. Tanguay,et al.  A quantitative HpaII-PCR assay to measure methylation of DNA from a small number of cells. , 1990, Nucleic acids research.

[5]  M. Becker‐André,et al.  Absolute mRNA quantification using the polymerase chain reaction (PCR). A novel approach by a PCR aided transcript titration assay (PATTY). , 1989, Nucleic acids research.

[6]  D. Y. Wu,et al.  The ligation amplification reaction (LAR)--amplification of specific DNA sequences using sequential rounds of template-dependent ligation. , 1989, Genomics.

[7]  B. K. Pal,et al.  Allele-specific enzymatic amplification of beta-globin genomic DNA for diagnosis of sickle cell anemia. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[8]  D. Y. Wu,et al.  Direct analysis of single nucleotide variation in human DNA and RNA using in situ dot hybridization. , 1989, DNA.

[9]  S. Perrin,et al.  8 – COMPETITIVE PCR FOR QUANTITATION OF mRNA , 1990 .

[10]  R. Myers,et al.  Detection and localization of single base changes by denaturing gradient gel electrophoresis. , 1987, Methods in enzymology.

[11]  A. Riggs,et al.  The nucleotide sequence of a cDNA clone containing the entire coding region for mouse X-chromosome-linked phosphoglycerate kinase. , 1986, Gene.