Plant disease resistance gene (R gene); defense response gene encode some conserved motifs. In the present work, a PCR strategy was used to clone resistance gene analogs (RGAs); defense gene analogs (DGAs) from Sea-is...Plant disease resistance gene (R gene); defense response gene encode some conserved motifs. In the present work, a PCR strategy was used to clone resistance gene analogs (RGAs); defense gene analogs (DGAs) from Sea-island cotton variety Hai7124 using oligonucleotide primers based on the nucleotide-binding site (NBS); serine/threonine kinase (STK) in the R-gene; pathogenesis-related proteins of class 2 (PR2) of defense response gene. 79 NBS sequences, 21 STK sequences; 11 DGAs were cloned from disease-resistance cotton. Phylogenic analysis of 79 NBS-RGAs; NBS-RGAs nucleotide sequences of cotton already deposited in GenBank identified one new sub-cluster. The deduced amino acid sequences of NBS-RGAs; STK-RGAs were divided into two distinct groups respectively: Toll/Interleukin-1 receptor (TIR) group; non-TIR group, A group; B group. The expression of RGAs; DGAs having consecutive open reading frame (ORF) was also investigated; it was found that 6 NBS-RGAs; 1 STK-RGA were induced,; 1 DGA was up-regulated by infection of Verticillium dahliae strain VD8. 4 TIR-NBS-RGAs; 4 non-TIR-NBS-RGAs were arbitrarily used as probes for Southern-blotting. There existed 2–10 blotted bands. In addition, since three non-TIR-NBS-RGAs have the same hybridization pattern, we conjecture that these three RGAs form a cluster distribution in the genome.展开更多
One resistance gene analog fragment named RGA-CIN14 was isolated from TcLr19 wheat,which contains kinase-2,kinase-3a,and the GLPL motif of the NBS-spanning region,using degenerated primers according to the nucleotide ...One resistance gene analog fragment named RGA-CIN14 was isolated from TcLr19 wheat,which contains kinase-2,kinase-3a,and the GLPL motif of the NBS-spanning region,using degenerated primers according to the nucleotide binding site （NBS） conserved domain.Based on the RGA-CIN14,a full-length cDNA,CIN14,which was 2 987 bp encoding 880 amino acids,was obtained by using the method of the rapid amplification cDNA ends （RACE）.Bioinformatics analysis showed that the deduced amino acids of CIN14 protein consisted of a NB-ARC conserved domain and many leucine-rich repeats （LRR） domains.The phylogenetic tree analysis indicated a considerable identity of the protein encoded by CIN14 with that of wheat leaf rust resistance gene Lr1,but a lower similarity with Lr21.The expression profile of the CIN14 gene detected by semi-quantitative RT-PCR showed that the CIN14 gene was not induced by Puccinia triticina and it was a constitutive gene with low abundance in the wheat leaf tissue.The resistance homology sequence was successfully obtained,which provides the shortcut for cloning of the resistance gene in TcLr19 wheat.展开更多
Disease-resistant genes play an important role in defending against a variety of pathogens and insect pests in plants. Most of the disease-resistant genes encode pro- teins with conserved leucine rich repeat and nucle...Disease-resistant genes play an important role in defending against a variety of pathogens and insect pests in plants. Most of the disease-resistant genes encode pro- teins with conserved leucine rich repeat and nucleotide binding site domains. In this study, we cloned and char- acterized gene-resistant analogs （RGAs） from Eucalyptus grandis using degenerate PCR, with primers specifically targeting these two domains. The amplified fragments were cloned into the pGEM-T vector and transformed into Escherichia coli. Among the 90 clones obtained, 13 were sequenced and compared with each other and with previ- ously identified gene-resistant diseases. A BLASTX search in GenBank revealed high similarities among the con- served domains of these cloned genes with RGA genes. Some clones, however, showed no significant similarity with DNA sequences in GenBank. Southern blotting ana- lysis identified several polymorphic RFLP loci between distinct genotypes. However, none of them co-segregated with the Puccinia psidii Winter resistance gene 1 （Ppr1） in a population study.展开更多
基金supported by the National Natural Science Foundation of China(Grant Nos.30270806 and 30370899)Program for Changjiang Scholars and Innovative Research Team in University in Ministry of Education in China.
文摘Plant disease resistance gene (R gene); defense response gene encode some conserved motifs. In the present work, a PCR strategy was used to clone resistance gene analogs (RGAs); defense gene analogs (DGAs) from Sea-island cotton variety Hai7124 using oligonucleotide primers based on the nucleotide-binding site (NBS); serine/threonine kinase (STK) in the R-gene; pathogenesis-related proteins of class 2 (PR2) of defense response gene. 79 NBS sequences, 21 STK sequences; 11 DGAs were cloned from disease-resistance cotton. Phylogenic analysis of 79 NBS-RGAs; NBS-RGAs nucleotide sequences of cotton already deposited in GenBank identified one new sub-cluster. The deduced amino acid sequences of NBS-RGAs; STK-RGAs were divided into two distinct groups respectively: Toll/Interleukin-1 receptor (TIR) group; non-TIR group, A group; B group. The expression of RGAs; DGAs having consecutive open reading frame (ORF) was also investigated; it was found that 6 NBS-RGAs; 1 STK-RGA were induced,; 1 DGA was up-regulated by infection of Verticillium dahliae strain VD8. 4 TIR-NBS-RGAs; 4 non-TIR-NBS-RGAs were arbitrarily used as probes for Southern-blotting. There existed 2–10 blotted bands. In addition, since three non-TIR-NBS-RGAs have the same hybridization pattern, we conjecture that these three RGAs form a cluster distribution in the genome.
基金funded by the National Natural Science Foundation of China (30771391,30700505)the Natural Science Foundation of Hebei Province,China (C2008000281)
文摘One resistance gene analog fragment named RGA-CIN14 was isolated from TcLr19 wheat,which contains kinase-2,kinase-3a,and the GLPL motif of the NBS-spanning region,using degenerated primers according to the nucleotide binding site （NBS） conserved domain.Based on the RGA-CIN14,a full-length cDNA,CIN14,which was 2 987 bp encoding 880 amino acids,was obtained by using the method of the rapid amplification cDNA ends （RACE）.Bioinformatics analysis showed that the deduced amino acids of CIN14 protein consisted of a NB-ARC conserved domain and many leucine-rich repeats （LRR） domains.The phylogenetic tree analysis indicated a considerable identity of the protein encoded by CIN14 with that of wheat leaf rust resistance gene Lr1,but a lower similarity with Lr21.The expression profile of the CIN14 gene detected by semi-quantitative RT-PCR showed that the CIN14 gene was not induced by Puccinia triticina and it was a constitutive gene with low abundance in the wheat leaf tissue.The resistance homology sequence was successfully obtained,which provides the shortcut for cloning of the resistance gene in TcLr19 wheat.
基金the Suzano Celulose S/A for logistical supportthe‘‘Conselho Nacional de Desenvolvimento Científico e Tecnológico’’(CNPq)+1 种基金the‘‘Coordenao de Aperfeioamento de Pessoal de Nível Superior’’(CAPES)the‘‘Fundao de Amparo à Pesquisa do Estado de Minas Gerais’’(FAPEMIG)for financial support to this study
文摘Disease-resistant genes play an important role in defending against a variety of pathogens and insect pests in plants. Most of the disease-resistant genes encode pro- teins with conserved leucine rich repeat and nucleotide binding site domains. In this study, we cloned and char- acterized gene-resistant analogs （RGAs） from Eucalyptus grandis using degenerate PCR, with primers specifically targeting these two domains. The amplified fragments were cloned into the pGEM-T vector and transformed into Escherichia coli. Among the 90 clones obtained, 13 were sequenced and compared with each other and with previ- ously identified gene-resistant diseases. A BLASTX search in GenBank revealed high similarities among the con- served domains of these cloned genes with RGA genes. Some clones, however, showed no significant similarity with DNA sequences in GenBank. Southern blotting ana- lysis identified several polymorphic RFLP loci between distinct genotypes. However, none of them co-segregated with the Puccinia psidii Winter resistance gene 1 （Ppr1） in a population study.
