Cloning of full-length methylmalonyl-CoA mutase from a cDNA library using the polymerase chain reaction.
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Jansen R, Kalousek F, Fenton WA, Rosenberg LE, Ledley FD
Cloning of full-length methylmalonyl-CoA mutase from a cDNA library using the polymerase chain reaction.
Genomics. 1989 Feb;4(2):198-205.
- PubMed ID
- 2567699 [ View in PubMed]
- Abstract
The polymerase chain reaction was used to clone a full-length human methylmalonyl-CoA mutase cDNA from a human liver library by priming with sequences from the 5' end of a partial cDNA and sequences in the phage vector. The amino acid sequence predicted from the cDNA corresponds to the authentic amino acid sequences of peptide fragment from purified methylmalonyl-CoA mutase. The open reading frame of the cDNA encodes 742 amino acids (82,283 Da) comprising a 32 amino acid mitochondrial leader sequence and a mature protein of 710 amino acids (78,489 Da). The use of the polymerase chain reaction to "screen" the cDNA library represents a novel application of this technique. The full length will enable analysis of mutations underlying inherited methylmalonic acidemias caused by deficiency of the methylmalonyl-CoA mutase apoenzyme.