Identification of a large cluster of coiled coil-nucleotide binding site--leucine rich repeat-type genes from the Rps1 region containing Phytophthora resistance genes in soybean.

Fifteen Rps genes confer resistance against the oomycete pathogen Phytophthora sojae, which causes root and stem rot disease in soybean. We have isolated a disease resistance gene-like sequence from the genomic region containing Rps1-k. Four classes of cDNA of the sequence were isolated from etiolated hypocotyl tissues that express the Rps1-k-encoded Phytophthora resistance. Sequence analyses of a cDNA clone showed that the sequence is a member of the coiled coil-nucleotide binding site-leucine rich repeat (CC-NBS-LRR)-type of disease resistance genes. It showed 36% identity to the recently cloned soybean resistance gene Rpg1-b, which confers resistance against Pseudomonas syringae pv. glycinea, and 56% and 38% sequence identity to putative resistance gene sequences from lotus and Medicago truncatula, respectively. The soybean genome contains about 38 copies of the sequence. Most of these copies are clustered in approximately 600 kb of contiguous DNA of the Rps1-k region. We have identified a recombinant that carries both rps1-k- and Rps1-k-haplotype-specific allelomorphs of two Rps1-k-linked molecular markers. An unequal crossover event presumably led to duplication of alleles for these two physically linked molecular markers. We hypothesize that the unequal crossing over was one of the mechanisms involved in tandem duplication of CC-NBS-LRR sequences in the Rps1-k region.

Bhattacharyya MK ，Narayanan NN ，Gao H ，Santra DK ，Salimath SS ，Kasuga T ，Liu Y ，Espinosa B ，Ellison L ，Marek L ，Shoemaker R ，Gijzen M ，Buzzell RI ... -  《THEORETICAL AND APPLIED GENETICS》

The soybean-Phytophthora resistance locus Rps1-k encompasses coiled coil-nucleotide binding-leucine rich repeat-like genes and repetitive sequences.

A series of Rps (resistance to Pytophthora sojae) genes have been protecting soybean from the root and stem rot disease caused by the Oomycete pathogen, Phytophthora sojae. Five Rps genes were mapped to the Rps1 locus located near the 28 cM map position on molecular linkage group N of the composite genetic soybean map. Among these five genes, Rps1-k was introgressed from the cultivar, Kingwa. Rps1-k has been providing stable and broad-spectrum Phytophthora resistance in the major soybean-producing regions of the United States. Rps1-k has been mapped and isolated. More than one functional Rps1-k gene was identified from the Rps1-k locus. The clustering feature at the Rps1-k locus might have facilitated the expansion of Rps1-k gene numbers and the generation of new recognition specificities. The Rps1-k region was sequenced to understand the possible evolutionary steps that shaped the generation of Phytophthora resistance genes in soybean. Here the analyses of sequences of three overlapping BAC clones containing the 184,111 bp Rps1-k region are reported. A shotgun sequencing strategy was applied in sequencing the BAC contig. Sequence analysis predicted a few full-length genes including two Rps1-k genes, Rps1-k-1 and Rps1-k-2. Previously reported Rps1-k-3 from this genomic region 1 was evolved through intramolecular recombination between Rps1-k-1 and Rps1-k-2 in Escherichia coli. The majority of the predicted genes are truncated and therefore most likely they are nonfunctional. A member of a highly abundant retroelement, SIRE1, was identified from the Rps1-k region. The Rps1-k region is primarily composed of repetitive sequences. Sixteen simple repeat and 63 tandem repeat sequences were identified from the locus. These data indicate that the Rps1 locus is located in a gene-poor region. The abundance of repetitive sequences in the Rps1-k region suggested that the location of this locus is in or near a heterochromatic region. Poor recombination frequencies combined with presence of two functional Rps genes at this locus has been providing stable Phytophthora resistance in soybean.

Gao H ，Bhattacharyya MK 《BMC PLANT BIOLOGY》

Two classes of highly similar coiled coil-nucleotide binding-leucine rich repeat genes isolated from the Rps1-k locus encode Phytophthora resistance in soybean.

A series of single genes protect soybean from the root and stem disease caused by the oomycete pathogen Phytophthora sojae. In the last two decades, Rps1-k has been the most stable and widely used Phytophthora resistance gene for the major soybean-producing regions of the United States. Four highly similar genes encoding coiled coil-nucleotide binding-leucine rich repeat (CC-NB-LRR)-type proteins were isolated from the Rps1-k locus. These genes were grouped into two classes based on their sequence identity. Class I contains three genes with identical open reading frames (ORF) and 5' end regions. Two of these genes were also identical at the 3' untranslated regions; the third gene showed a recombination breakpoint in the 3' untranslated region resulting in the combination of 3' end sequences of members from both classes. Reverse transcription-polymerase chain reaction analyses suggested that members of both classes of genes are transcribed at low levels. Representative members from each gene class were expressed in transgenic soybean plants. Analyses of independent R0, R1, R2, and R3 progeny populations suggested that both gene classes confer Phytophthora resistance in soybean. A possible evolutionary mechanism for the Class I gene family is proposed.

Gao H ，Narayanan NN ，Ellison L ，Bhattacharyya MK ... -  《MOLECULAR PLANT-MICROBE INTERACTIONS》

Identification and molecular mapping of Rps11, a novel gene conferring resistance to Phytophthora sojae in soybean.

Rps11 confers excellent resistance to predominant Phytophthora sojae isolates capable of defeating major Rps genes deployed into soybean production, representing a novel source of resistance for soybean cultivar enhancement. Phytophthora root and stem rot (PRSR), caused by the soil-borne pathogen Phytophthora sojae, is a devastating disease of soybean [Glycine max (L.) Merr.] throughout the world. Deploying resistant soybean cultivars is the most effective and environmentally friendly approach to managing this disease. The soybean landrace PI 594527 was found to carry excellent resistance to all P. sojae isolates examined, some of which were capable of overcoming the major Rps genesp, such as Rps1-k, Rps1-c, and Rps3-a, predominantly used for soybean protection in the past decades. A mapping population consisting of 58 F2 individuals and 209 F2:3 families derived from a cross between PI 594527 and the susceptible cultivar 'Williams' was used to characterize the inheritance pattern of the resistance to P. soja (Rps) in PI 594527. It was found that the resistance was conferred by a single Rps gene, designated Rps11, which was initially defined as an ~5 Mb genomic region at the beginning of chromosome 7 by bulked segregant analysis (BSA) with a nucleotide polymorphism (SNP) chip comprising 7039 SNP markers. Subsequently, simple sequence repeat (SSR) markers in the defined region were used to genotype the F2:3 mapping population to map Rps11 to a 225.3 kb genomic region flanked by SSR markers BARCSOYSSR_07_0286 and BARCSOYSSR_07_0300, according to the soybean reference genome sequence. Particularly, an SSR marker (i.e., BARCSOYSSR_07_0295) was found to tightly co-segregate with Rps11 in the mapping population and can be effectively used for marker-assisted selection of this gene for development of resistant soybean cultivars.

Ping J ，Fitzgerald JC ，Zhang C ，Lin F ，Bai Y ，Wang D ，Aggarwal R ，Rehman M ，Crasta O ，Ma J ... -  《-》

A copia-like retrotransposon Tgmr closely linked to the Rps1-k allele that confers race-specific resistance of soybean to Phytophthora sojae.

We have isolated and characterized Tgmr, a copia-like retrotransposon, linked tightly to the Rps1-k allele that confers race-specific resistance of soybean to the the fungal pathogen Phytophthora sojae. Southern analysis followed by PCR and sequence analyses, using primers based on sequences flanking the insertion site confirmed that the element was inserted in the neighboring region of Rps1-k but not in that of the other four Rps1 alleles. This implies that Tgmr was transposed into the Rps1-k flanking site after the divergence of Rps1 alleles. Southern analysis of a series of diverse soybean cultivars revealed a high level of polymorphism of Tgmr-related sequences. These results indicate that this low copy retroelement family could have been active in the soybean genome in the recent past. Tgmr contains long terminal repeats (LTR) and four non-overlapping open reading frames (ORF), presumably originating from mutations leading to stop codons of a single ORF. The conserved domains for gag, protease, integrase, reverse transcriptase and RNaseH are present in the internal portion of the element. However, the protease, reverse transcriptase and RNaseH of this element are non-functional due to the presence of several stop codons. Possible transactivation of Tgmr and application of this element in insertional mutagenesis for soybean are discussed.

Bhattacharyya MK ，Gonzales RA ，Kraft M ，Buzzell RI ... -  《PLANT MOLECULAR BIOLOGY》