scholarly journals Isolation, Characterization and Phylogenetic Analysis of Nucleotide Binding Site-encoding Disease-resistance Gene Analogues from European Aspen (Populus tremula)

2010 ◽  
Vol 59 (1-6) ◽  
pp. 68-77 ◽  
Author(s):  
Yong Zhang ◽  
Shougong Zhang ◽  
Liwang Qi ◽  
Tao Zhang ◽  
Chunguo Wang ◽  
...  
Keyword(s):  
Disease Resistance ◽  
Binding Site ◽  
Resistance Gene ◽  
Nucleotide Binding ◽  
Nucleotide Binding Site ◽  
Populus Tremula ◽  
R Gene ◽  
R Genes ◽  
Degenerate Primers ◽  
European Aspen

Abstract The majority of verified plant disease resistance genes (R genes) isolated to date was of the nucleotide binding site-leucine rich repeat (NBS-LRR) class. The conservation between different NBS-LRR R genes opens the avenue for the use of PCR based strategies in isolating and cloning other R gene family members or analogs (resistance gene analogue, RGA) using degenerate primers for these conserved regions. In this study, to better understand the R gene in European aspen (Populus tremula), a perennial tree, we used degenerate primers to amplify RGA sequences from European aspen. Cloning and sequence characterization identified 37 European aspen RGAs, which could be phylogenetically classified into seven subfamilies. Deduced amino acid sequences of European aspen RGAs showed strong identity, ranging from 30.41 to 46.63%, to toll interleukin receptor (TIR) R gene subfamily. BLAST searches with reference to the genomic sequence of P. trichocarpa found 209 highly homologous regions distributed in 28 genomic loci, suggesting the abundance and divergence of NBS-encoding R genes in European aspen genome. Although, numerous studies have reported that plant R genes are under diversifying selection for specificity to evolving pathogens, non-synonymous to synonymous nucleotide substitution (dN/dS) ratio were <1 for NBS domains of European aspen RGA, showing the evidence of purifying selection in this perennial tree. In further analysis, many intergenic exchanges were also detected among these RGAs, indicating a probable role in homogenising NBS domains. The present study permits insights into the origin, diversification, evolution and function of NBS-LRR R genes in perennial species like European aspen and will be useful for further R gene isolation and exploitation.

scholarly journals Isolation and Characterization of Nucleotide-Binding Site Resistance Gene Homologues in Common Bean (Phaseolus vulgaris)

2013 ◽  
Vol 103 (2) ◽  
pp. 156-168 ◽  
Author(s):  
Luz N. Garzón ◽  
Oscar A. Oliveros ◽  
Benjamin Rosen ◽  
Gustavo A. Ligarreto ◽  
Douglas R. Cook ◽  
...  
Keyword(s):  
Common Bean ◽  
Binding Site ◽  
Resistance Gene ◽  
Focal Point ◽  
Nucleotide Binding ◽  
Nucleotide Binding Site ◽  
R Genes ◽  
Degenerate Primers ◽  
Model Legume ◽  
Isolation And Characterization

Common bean production is constrained by many fungal, viral, and bacterial pathogens. Thus, the identification of resistance (R) genes is an important focal point of common bean research. The main goal of our study was to identify resistance gene homologues (RGH) in the crop, using degenerate primers designed from conserved sequences in the nucleotide-binding site (NBS) domains of R-genes from the model legume Medicago truncatula. Total DNA of the Andean common bean genotype G19833 was used for amplification of over 500 primer combinations. Sequencing of amplicons showed that 403 cloned fragments had uninterrupted open reading frames and were considered representative of functional RGH genes. The sequences were grouped at two levels of nucleotide identity (90 and 80%) and representative sequences of each group were used for phylogenetic analyses. The RGH sequence diversity of common bean was divided into TIR and non-TIR families, each with different clusters. The TIR sequences grouped into 14 clades while non-TIR sequences grouped into seven clades. Pairwise comparisons showed purifying selection, although some sequences may have been the result of diversifying selection. Knowledge about RGH genes in common bean can allow the design of molecular markers for pyramiding of resistance genes against various pathogens.

Isolation of a family of resistance gene analogue sequences of the nucleotide binding site (NBS) type fromLensspecies

Genome ◽  
2004 ◽  
Vol 47 (4) ◽  
pp. 650-659 ◽  
Author(s):  
M W.F Yaish ◽  
L E Sáenz de Miera ◽  
M Pérez de la Vega
Keyword(s):  
Disease Resistance ◽  
Binding Site ◽  
Resistance Gene ◽  
Resistance Genes ◽  
Nucleotide Binding ◽  
Nucleotide Binding Site ◽  
Disease Resistance Genes ◽  
Nucleotide Sequence Analysis ◽  
Degenerate Primers ◽  
Conserved Motifs

Most known plant disease-resistance genes (R genes) include in their encoded products domains such as a nucleotide-binding site (NBS) or leucine-rich repeats (LRRs). Sequences with unknown function, but encoding these conserved domains, have been defined as resistance gene analogues (RGAs). The conserved motifs within plant NBS domains make it possible to use degenerate primers and PCR to isolate RGAs. We used degenerate primers deduced from conserved motifs in the NBS domain of NBS-LRR resistance proteins to amplify genomic sequences from Lens species. Fragments from approximately 500-850 bp were obtained. The nucleotide sequence analysis of these fragments revealed 32 different RGA sequences in Lens species with a high similarity (up to 91%) to RGAs from other plants. The predicted amino acid sequences showed that lentil sequences contain all the conserved motifs (P-loop, kinase-2, kinase-3a, GLPL, and MHD) present in the majority of other known plant NBS–LRR resistance genes. Phylogenetic analyses grouped the Lens NBS sequences with the Toll and interleukin-1 receptor (TIR) subclass of NBS–LRR genes, as well as with RGA sequences isolated from other legume species. Using inverse PCR on one putative RGA of lentil, we were able to amplify the flanking regions of this sequence, which contained features found in R proteins.Key words: disease resistance genes, comparative analysis, lentils, TIR, LRR.

A superfamily of disease resistance gene analogs is located on all homoeologous chromosome groups of wheat (Triticum aestivum)

Genome ◽  
1998 ◽  
Vol 41 (6) ◽  
pp. 782-788 ◽  
Author(s):  
W Spielmeyer ◽  
M Robertson ◽  
N Collins ◽  
D Leister ◽  
P Schulze-Lefert ◽  
...  
Keyword(s):  
Disease Resistance ◽  
Binding Site ◽  
Resistance Gene ◽  
Resistance Genes ◽  
Nucleotide Binding ◽  
Nucleotide Binding Site ◽  
Resistance Gene Analogs ◽  
Leucine Rich Repeat ◽  
Homoeologous Chromosome ◽  
Entry Points

In this study, resistance gene analogs (RGAs) which were isolated from monocot crop species (wheat, barley, maize and rice) and contained conserved sequence motifs found within the nucleotide binding site - leucine rich repeat (NBS-LRR) class of resistance genes, were used to assess their distribution in the wheat genome. The RGAs showed 30-70% amino acid identity to a previously isolated monocot NBS-LRR sequence from the Cre3 locus for cereal cyst nematode (CCN) resistance in wheat. We used the RGAs as probes to identify and map loci in wheat using recombinant inbred lines of an international Triticeae mapping family. RGA loci mapped across all seven homoeologous chromosome groups of wheat. This study demonstrated that the RGA mapping approach provides potential entry points toward identifying resistance gene candidates in wheat.Key words: wheat, disease resistance genes, nucleotide binding site, leucine rich repeat, resistance gene analogs.

Candidate disease resistance genes in sunflower cloned using conserved nucleotide-binding site motifs: Genetic mapping and linkage to the downy mildew resistance gene Pl1

Genome ◽  
2001 ◽  
Vol 44 (2) ◽  
pp. 205-212 ◽  
Author(s):  
Melaku Ayele Gedil ◽  
Mary B Slabaugh ◽  
Simon Berry ◽  
Richard Johnson ◽  
Richard Michelmore ◽  
...  
Keyword(s):  
Disease Resistance ◽  
Binding Site ◽  
Downy Mildew ◽  
Resistance Gene ◽  
Nucleotide Binding ◽  
Nucleotide Binding Site ◽  
Sequence Motifs ◽  
Caps Marker ◽  
Race 1 ◽  
Dna Sequence Motifs

Disease resistance gene candidates (RGCs) belonging to the nucleotide-binding site (NBS) superfamily have been cloned from numerous crop plants using highly conserved DNA sequence motifs. The aims of this research were to (i) isolate genomic DNA clones for RGCs in cultivated sunflower (Helianthus annuus L.) and (ii) map RGC markers and Pl1, a gene for resistance to downy mildew (Plasmopara halstedii (Farl.) Berl. & de Toni) race 1. Degenerate oligonucleotide primers targeted to conserved NBS DNA sequence motifs were used to amplify RGC fragments from sunflower genomic DNA. PCR products were cloned, sequenced, and assigned to 11 groups. RFLP analyses mapped six RGC loci to three linkage groups. One of the RGCs (Ha-4W2) was linked to Pl1, a downy mildew resistance gene. A cleaved amplified polymorphic sequence (CAPS) marker was developed for Ha-4W2 using gene-specific oligonucleotide primers. Downy mildew susceptible lines (HA89 and HA372) lacked a 276-bp Tsp509I restriction fragment that was present in downy mildew resistant lines (HA370, 335, 336, 337, 338, and 339). HA370 × HA372 F2 progeny were genotyped for the Ha-4W2 CAPS marker and phenotyped for resistance to downy mildew race 1. The CAPS marker was linked to but did not completely cosegregate with Pl1 on linkage group 8. Ha-4W2 was found to comprise a gene family with at least five members. Although genetic markers for Ha-4W2 have utility for marker-assisted selection, the RGC detected by the CAPS marker has been ruled out as a candidate gene for Pl1. Three of the RGC probes were monomorphic between HA370 and HA372 and still need to be mapped and screened for linkage to disease resistance loci.Key words: Helianthus, sunflower, downy mildew, Plasmopara, nucleotide-binding site.

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