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Updated: May 19, 2026

Merging Absolute and Relative Quantitative PCR Data to Quantify STAT3 Splice Variant Transcripts
Published on: October 9, 2016
The exon 29 c.3535A>T in the alpha-2-macroglobulin gene causing aberrant splice variants is associated with mastitis
Xiuge Wang1, Jinming Huang, Lihong Zhao
1Laboratory of Molecular Genetics and Breeding, Center of Dairy Cattle Research, Shandong Academy of Agricultural Sciences, Industry North Road 159, Jinan 250131, Shandong, People's Republic of China.
Abstract:
Alpha-2-macroglobulin (A2M) binds proteases, thereby acting as defense barriers against pathogens in the plasma and tissues of vertebrates and invertebrates. Quantitative real-time polymerase chain reaction (PCR) and the isobaric tags for relative and absolute quantitation method were used to determine the expression levels of A2M mRNA and proteins in mastitis-infected mammary tissues. A2M mRNA and protein expression were significantly higher in mastitis-infected mammary tissues than those in healthy tissues. We also identified 23 novel A2M splice variants in the bovine mammary tissues using reverse transcription PCR combined with clone sequencing. These splice variants predominantly affected the bait region, the inhibitory region, and the thioester region of the protein, which have the functional key roles in inhibiting the proteases of pathogens. Genomic sequencing analysis revealed a nonsynonymous c.3535A>T single-nucleotide polymorphism (SNP) in exon 29, which is located within a putative exonic splice enhancer and may be the reason why the A2M gene produces the aberrant splice variant A2M-AS4. Our findings suggest that the A2M gene can play its role by alternative splicing mechanism and it may be of significance against mastitis. This study provides clues to better understand the function of the bovine A2M gene and the effects of the exonic SNP on the production of aberrant splice variants.
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