Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance genes in lettuce

Katherine A. Shen, Blake C. Meyers, M. Nurul Islam-Faridi, Doris B. Chin, David M. Stelly, Richard W Michelmore

Research output: Contribution to journalArticle

190 Citations (Scopus)

Abstract

The recent cloning of genes for resistance against diverse pathogens from a variety of plants has revealed that many share conserved sequence motifs. This provides the possibility of isolating numerous additional resistance genes by polymerase chain reaction (PCR) with degenerate oligonucleotide primers. We amplified resistance gene candidates (RGCs) from lettuce with multiple combinations of primers with low degeneracy designed from motifs in the nucleotide binding sites (NBSs) of RPS2 of Arabidopsis thaliana and N of tobacco. Genomic DNA, cDNA, and bacterial artificial chromosome (BAC) clones were successfully used as templates. Four families of sequences were identified that had the same similarity to each other as to resistance genes from other species. The relationship of the amplified products to resistance genes was evaluated by several sequence and genetic criteria. The amplified products contained open reading frames with additional sequences characteristic of NBSs. Hybridization of RGCs to genomic DNA and to BAC clones revealed large numbers of related sequences. Genetic analysis demonstrated the existence of clustered multigene families for each of the four RGC sequences. This parallels classical genetic data on clustering of disease resistance genes. Two of the four families mapped to known dusters of resistance genes; these two families were therefore studied in greater detail. Additional evidence that these RGCs could be resistance genes was gained by the identification of leucine-rich repeat (LRR) regions in sequences adjoining the NBS similar to those in RPM1 and RPS2 of A. thaliana. Fluorescent in situ hybridization confirmed the clustered genomic distribution of these sequences. The use of PCR with degenerate oligonucleotide primers is therefore an efficient method to identify numerous RGCs in plants.

Original languageEnglish (US)
Pages (from-to)815-823
Number of pages9
JournalMolecular Plant-Microbe Interactions
Volume11
Issue number8
StatePublished - Aug 1998

Fingerprint

Lettuce
DNA Primers
Polymerase chain reaction
DNA primers
Multigene Family
lettuce
Genes
polymerase chain reaction
Polymerase Chain Reaction
genes
binding sites
bacterial artificial chromosomes
Bacterial Artificial Chromosomes
genomics
Nucleotides
Binding Sites
Chromosomes
Arabidopsis
Arabidopsis thaliana
clones

ASJC Scopus subject areas

  • Microbiology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biotechnology

Cite this

Shen, K. A., Meyers, B. C., Islam-Faridi, M. N., Chin, D. B., Stelly, D. M., & Michelmore, R. W. (1998). Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance genes in lettuce. Molecular Plant-Microbe Interactions, 11(8), 815-823.

Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance genes in lettuce. / Shen, Katherine A.; Meyers, Blake C.; Islam-Faridi, M. Nurul; Chin, Doris B.; Stelly, David M.; Michelmore, Richard W.

In: Molecular Plant-Microbe Interactions, Vol. 11, No. 8, 08.1998, p. 815-823.

Research output: Contribution to journalArticle

Shen, KA, Meyers, BC, Islam-Faridi, MN, Chin, DB, Stelly, DM & Michelmore, RW 1998, 'Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance genes in lettuce', Molecular Plant-Microbe Interactions, vol. 11, no. 8, pp. 815-823.
Shen, Katherine A. ; Meyers, Blake C. ; Islam-Faridi, M. Nurul ; Chin, Doris B. ; Stelly, David M. ; Michelmore, Richard W. / Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance genes in lettuce. In: Molecular Plant-Microbe Interactions. 1998 ; Vol. 11, No. 8. pp. 815-823.
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