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Consensus-degenerate hybrid oligonucleotide primers for amplification of distantly related sequences.
Title | Consensus-degenerate hybrid oligonucleotide primers for amplification of distantly related sequences. |
Publication Type | Journal Article |
Year of Publication | 1998 |
Authors | Rose, TM, Schultz, ER, Henikoff, JG, Pietrokovski, S, McCallum, CM, Henikoff, S |
Journal | Nucleic Acids Res |
Volume | 26 |
Issue | 7 |
Pagination | 1628-35 |
Date Published | 1998 Apr 1 |
ISSN | 0305-1048 |
Keywords | Amino Acid Sequence, Animals, Arthritis, Rheumatoid, Base Sequence, Codon, Computer Communication Networks, Consensus Sequence, Conserved Sequence, DNA Modification Methylases, DNA Primers, Evolution, Molecular, Humans, Molecular Sequence Data, Nucleic Acid Hybridization, Phylogeny, Polymerase Chain Reaction, RNA-Directed DNA Polymerase, Sarcoma, Kaposi, Sequence Alignment, Sequence Homology, Amino Acid, Sequence Homology, Nucleic Acid, Software |
Abstract | We describe a new primer design strategy for PCR amplification of unknown targets that are related to multiply-aligned protein sequences. Each primer consists of a short 3' degenerate core region and a longer 5' consensus clamp region. Only 3-4 highly conserved amino acid residues are necessary for design of the core, which is stabilized by the clamp during annealing to template molecules. During later rounds of amplification, the non-degenerate clamp permits stable annealing to product molecules. We demonstrate the practical utility of this hybrid primer method by detection of diverse reverse transcriptase-like genes in a human genome, and by detection of C5DNA methyltransferase homologs in various plant DNAs. In each case, amplified products were sufficiently pure to be cloned without gel fractionation. This COnsensus-DEgenerate Hybrid Oligonucleotide Primer (CODEHOP) strategy has been implemented as a computer program that is accessible over the World Wide Web (http://blocks.fhcrc.org/codehop.html) and is directly linked from the BlockMaker multiple sequence alignment site for hybrid primer prediction beginning with a set of related protein sequences. |
Alternate Journal | Nucleic Acids Res. |
PubMed ID | 9512532 |
PubMed Central ID | PMC147464 |