Sep 1, 1989

Porcine tumor necrosis factor alpha: cloning with the polymerase chain reaction and determination of the nucleotide sequence

U PauliE Peterhans


We have cloned the gene for the porcine tumor necrosis factor alpha (TNF-alpha) utilizing the polymerase chain reaction (PCR). Total RNA from stimulated monocytes was used to generate TNF-alpha-specific single-stranded cDNA, with reverse transcriptase and a conserved consensus primer, starting at the TNF-alpha translation stop codon. After adding a second conserved consensus primer to the propeptide region, we amplified the cDNA between the two primers. The isolated fragment was cloned and sequenced. Comparison of the nucleotide sequence indicated an 85% sequence similarity to the human TNF-alpha gene. Eighteen amino acids of the deduced mature peptide sequence of porcine TNF-alpha were different from those in the sequence of man. The technique used in this work allows rapid cloning of specific genes from total RNA, and, additionally, screens for full-length transcripts during the amplification procedure.

Mentioned in this Paper

Tumor Necrosis Factor-alpha
Electrophoresis, Agar Gel
Gene Amplification
Amino Acids, I.V. solution additive
Codon, Terminator
Polymerase Chain Reaction Analysis
Gene Amplification Technique
Peptide Fragments

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