Sheshu Madhav - Academia.edu (original) (raw)
Papers by Sheshu Madhav
Molecular Breeding, 2011
Rice blast is one of the most devastating diseases affecting the rice crop throughout the world. ... more Rice blast is one of the most devastating diseases affecting the rice crop throughout the world. In molecular breeding for host plant resistance, functional markers are very useful for enhancing the precision and accuracy in marker-assisted selection (MAS) of target gene(s) with minimum effort, time and cost. Pi54 (which was earlier known as Pik h ) is one of the major blast resistance genes and has been observed to show resistance against many isolates of the blast pathogen in India. The gene has been cloned through map-based strategy and encodes a nucleotide-binding site–leucine-rich repeat (NBS–LRR) domain-containing protein. In the present study, we carried out allele mining for this gene and identified a 144-bp insertion/deletion (InDel) polymorphism in the exonic region of the gene. A PCR-based co-dominant molecular marker targeting this InDel, named Pi54 MAS, was developed. Pi54 MAS was observed to perfectly co-segregate with blast resistance in a mapping population with no recombinants. Validation of this marker in 105 genotypes which are either susceptible or resistant to rice blast disease showed that the marker is polymorphic in most of the resistant–susceptible genotype combinations and is more accurate than the earlier reported markers for Pi54. Hence this functional, co-dominant marker is suggested for routine deployment in MAS of Pi54 in breeding programs.
Rice, 2010
The beet armyworm (Spodoptera exigua) and the rice water weevil (Lissorhoptrus oryzophilus) are t... more The beet armyworm (Spodoptera exigua) and the rice water weevil (Lissorhoptrus oryzophilus) are two important insect pests in rice production. To identify insect-responsive genes in rice, we performed a deep transcriptome analysis of Nipponbare rice leaves infested with both beet armyworm and water weevil using massively parallel signature sequencing (MPSS). Many antisense, alternative, and novel transcripts were commonly and specifically induced and suppressed in the infested tissue. Key genes involved in the defense metabolic pathways such as salicylic acid and jasmonic acid biosynthesis pathways were up-regulated in the infested leaves. To validate the MPSS results, we analyzed the transcriptome of the rice leaves infested with water weevils using Solexa’s sequencing-by-synthesis (SBS) method. The MPSS and SBS data were highly correlated (Pearson’s correlation coefficient = 0.85), and 83% of genes had similar gene expression in both libraries. Our comprehensive and in-depth survey of the insect-infested libraries provides a rich genomic resource for further analyzing the function of key regulatory genes involved in insect resistance in rice.
Journal of Phytopathology, 2009
Identification of full length genes along with upstream regulatory elements is important to under... more Identification of full length genes along with upstream regulatory elements is important to understand its expression. Here, we report preparation of high titre genomic library and identification of a genomic clone containing Pi-kh gene with its complete upstream and downstream sequences from the rice blast resistant line Tetep. Structural analysis of protein revealed that Pi-kh has a central nucleotide binding site domain, leucine-rich repeats domain and a unique zinc-finger domain. Comparative analysis of Pi-kh protein sequence showed 64% and 45% similarity with the protein sequences of rice blast resistance genes Pi-b and Pi-ta, respectively.
Molecular Breeding, 2010
Kernel length in rice (Oryza sativa L.) is controlled by various quantitative trait loci of which... more Kernel length in rice (Oryza sativa L.) is controlled by various quantitative trait loci of which GS3 is the most important, being responsible for 80–90% of the variation in kernel length. A mutation in the second exon of this gene has been reported to be associated with maximum variations in the kernel length. We have developed a simple PCR-based marker system named DRR-GL which targets the functional nucleotide polymorphism at GS3. This marker system has the advantages that it is easy to use, saves time and cost, and is amenable for large-scale marker-assisted selection for the trait of kernel length. Validation of this marker in a segregating population and 152 rice varieties, which includes 30 elite basmati varieties, reveals its effective co-segregation and association with the traits of kernel length as well as kernel elongation after cooking. We recommend utilization of this simple, low-cost marker system in breeding programs targeted at improvement of key rice grain quality traits, kernel length and kernel elongation.
Molecular Breeding
Depending on the position on the panicle, grain filling differs for spikelets on primary branches... more Depending on the position on the panicle, grain filling differs for spikelets on primary branches and secondary branches of upper and lower portions in rice (Oryza sativa L.). With the objective of identifying candidate genes associated with the grain-filling process across the panicle, a candidate gene (CG)-based mapping approach was attempted in an F 2 population derived from the cross between Rasi, a rice variety known for its good grain filling, and IC114927, a local landrace. The grain filling (%) was characterized for 444 F 2 individual plants across the panicle. Within the F 2 population, thirteen plants were identified with >90% grain filling across the panicle. For 120 markers designed from 1 kb upstream and within the candidate genes reported to be involved in translocation process of sugars, 18.3% polymorphism was observed. Using a selective genotyping approach comprising 24 individuals from eight groups based on their filling and position, two associated CG markers were identified. Further analysis of the mapping population showed a CG marker based on the sucrose phosphate synthase gene on chromosome 2 to be significantly associated with filling of grains on primary branches of the upper half of the panicle and another CG marker based on the transporter gene on chromosome 11 to be associated with filling of grains on primary branches of the lower half; secondary branches of the upper half and lower half of the panicle. Both the positive alleles were contributed by the variety Rasi. With a candidate gene-based marker mapping approach, two genes involved in sugar translocation and transport were identified to be directly associated with grain filling in rice.
During the past five decades, a large number of tobacco varieties have been developed for differe... more During the past five decades, a large number of tobacco varieties have been developed for different end uses in India through pure line selection from local land-races, mutation breeding and hybridization involving local selections and exotic introductions followed by pedi- gree selection. No systematic effort has been made to understand the existing diversity pattern in these va- rieties, which is
Euphytica, 2009
SSR markers targeting (GATA) n motifs are known to be highly polymorphic and useful in many organ... more SSR markers targeting (GATA) n motifs are known to be highly polymorphic and useful in many organisms. (GATA) n motif specific SSR markers covering the whole rice genome are not available. The present study was carried out with an objective to identify class I rice microsatellites in the rice genome with (GATA) n motifs, in-silico, and validate their potential as molecular markers. A total of 243 such motifs were identified; 65 of these were present in the genic region, 59 were in the upstream region and the remaining motifs were found in the intergenic regions. Many of the (GATA) n motifs were found within and/or upstream of genes associated with biotic or abiotic stress tolerance. A total of 230 PCRbased markers targeting all the class I (GATA) n microsatellites were developed and 35 of these markers spread across the rice genome were validated in a set of 24 representative rice varieties belonging to five distinct cultivar groups. All the markers were polymorphic, with average polymorphism information content (PIC) value of 0.61, and the rice cultivars could be uniquely distinguished into different cultivar groups based on marker analysis. These informative markers targeting (GATA) n motifs representing a new set of markers in rice will be highly useful for genetic studies and marker-assisted selection.
Proceedings of 2nd …, 2006
Molecular …, 2009
Fragrance development in rice has been reported due to a 8-bp deletion in the exon 7 of badh2 gen... more Fragrance development in rice has been reported due to a 8-bp deletion in the exon 7 of badh2 gene located on Chromosome 8S. Multiplex markers targeting the functional InDel polymorphism was earlier reported for genotyping fragrance trait, but the marker was observed to be inconsistent and difficult to use. We have developed a simple, co-dominant, functional marker for fragrance trait, which can be resolved in an agarose gel and validated in Basmati and non-Basmati aromatic rice varieties and in a mapping population segregated for fragrance trait. The marker targets the InDel polymorphism in badh2 gene and amplifies 95 and 103 bp fragments in fragrant and non-fragrant genotypes, respectively. The newly developed marker was highly efficient in discriminating all fragrant and non-fragrant genotypes and showed perfect co-segregation with the trait of fragrance in the mapping population. We recommend the use of this simple, low-cost marker in routine genotyping for fragrance trait in large scale breeding materials and germplasm.
Biotechnology Advances, 2010
Enormous sequence information is available in public databases as a result of sequencing of diver... more Enormous sequence information is available in public databases as a result of sequencing of diverse crop genomes. It is important to use this genomic information for the identification and isolation of novel and superior alleles of agronomically important genes from crop gene pools to suitably deploy for the development of improved cultivars. Allele mining is a promising approach to dissect naturally occurring allelic variation at candidate genes controlling key agronomic traits which has potential applications in crop improvement programs. It helps in tracing the evolution of alleles, identification of new haplotypes and development of allele-specific markers for use in marker-assisted selection. Realizing the immense potential of allele mining, concerted allele mining efforts are underway in many international crop research institutes. This review examines the concepts, approaches and applications of allele mining along with the challenges associated while emphasizing the need for more refined ‘mining’ strategies for accelerating the process of allele discovery and its utilization in molecular breeding.
Molecular Genetics and Genomics, 2005
In order to understand the molecular mechanisms involved in the gene-for-gene type of pathogen re... more In order to understand the molecular mechanisms involved in the gene-for-gene type of pathogen resistance, high-resolution genetic and physical mapping of resistance loci is required to facilitate map-based cloning of resistance genes. Here, we report the molecular mapping and cloning of a dominant gene (Pi-k h ) present in the rice line Tetep, which is associated with resistance to rice blast disease caused by Magnaporthe grisea. This gene is effective against M. grisea populations prevalent in the Northwestern Himalayan region of India. Using 178 sequence tagged microsatellite, sequence-tagged site, expressed sequence tag and simple sequence repeat (SSR) markers to genotype a population of 208 F2 individuals, we mapped the Pi-k h gene between two SSR markers (TRS26 and TRS33) which are 0.7 and 0.5 cM away, respectively, and can be used in marker-assisted-selection for blast-resistant rice cultivars. We used the markers to identify the homologous region in the genomic sequence of Oryza sativa cv. Nipponbare, and a physical map consisting of two overlapping bacterial artificial chromosome and P1 artificial chromosome clones was assembled, spanning a region of 143,537 bp on the long arm of chromosome 11. Using bioinformatic analyses, we then identified a candidate blast-resistance gene in the region, and cloned the homologous sequence from Tetep. The putative Pi-k h gene cloned from Tetep is 1.5 kbp long with a single ORF, and belongs to the nucleotide binding site-leucine rich repeat class of disease resistance genes. Structural and expression analysis of the Pi-k h gene revealed that its expression is pathogen inducible.
Molecular Breeding, 2011
Rice blast is one of the most devastating diseases affecting the rice crop throughout the world. ... more Rice blast is one of the most devastating diseases affecting the rice crop throughout the world. In molecular breeding for host plant resistance, functional markers are very useful for enhancing the precision and accuracy in marker-assisted selection (MAS) of target gene(s) with minimum effort, time and cost. Pi54 (which was earlier known as Pik h ) is one of the major blast resistance genes and has been observed to show resistance against many isolates of the blast pathogen in India. The gene has been cloned through map-based strategy and encodes a nucleotide-binding site–leucine-rich repeat (NBS–LRR) domain-containing protein. In the present study, we carried out allele mining for this gene and identified a 144-bp insertion/deletion (InDel) polymorphism in the exonic region of the gene. A PCR-based co-dominant molecular marker targeting this InDel, named Pi54 MAS, was developed. Pi54 MAS was observed to perfectly co-segregate with blast resistance in a mapping population with no recombinants. Validation of this marker in 105 genotypes which are either susceptible or resistant to rice blast disease showed that the marker is polymorphic in most of the resistant–susceptible genotype combinations and is more accurate than the earlier reported markers for Pi54. Hence this functional, co-dominant marker is suggested for routine deployment in MAS of Pi54 in breeding programs.
Rice, 2010
The beet armyworm (Spodoptera exigua) and the rice water weevil (Lissorhoptrus oryzophilus) are t... more The beet armyworm (Spodoptera exigua) and the rice water weevil (Lissorhoptrus oryzophilus) are two important insect pests in rice production. To identify insect-responsive genes in rice, we performed a deep transcriptome analysis of Nipponbare rice leaves infested with both beet armyworm and water weevil using massively parallel signature sequencing (MPSS). Many antisense, alternative, and novel transcripts were commonly and specifically induced and suppressed in the infested tissue. Key genes involved in the defense metabolic pathways such as salicylic acid and jasmonic acid biosynthesis pathways were up-regulated in the infested leaves. To validate the MPSS results, we analyzed the transcriptome of the rice leaves infested with water weevils using Solexa’s sequencing-by-synthesis (SBS) method. The MPSS and SBS data were highly correlated (Pearson’s correlation coefficient = 0.85), and 83% of genes had similar gene expression in both libraries. Our comprehensive and in-depth survey of the insect-infested libraries provides a rich genomic resource for further analyzing the function of key regulatory genes involved in insect resistance in rice.
Journal of Phytopathology, 2009
Identification of full length genes along with upstream regulatory elements is important to under... more Identification of full length genes along with upstream regulatory elements is important to understand its expression. Here, we report preparation of high titre genomic library and identification of a genomic clone containing Pi-kh gene with its complete upstream and downstream sequences from the rice blast resistant line Tetep. Structural analysis of protein revealed that Pi-kh has a central nucleotide binding site domain, leucine-rich repeats domain and a unique zinc-finger domain. Comparative analysis of Pi-kh protein sequence showed 64% and 45% similarity with the protein sequences of rice blast resistance genes Pi-b and Pi-ta, respectively.
Molecular Breeding, 2010
Kernel length in rice (Oryza sativa L.) is controlled by various quantitative trait loci of which... more Kernel length in rice (Oryza sativa L.) is controlled by various quantitative trait loci of which GS3 is the most important, being responsible for 80–90% of the variation in kernel length. A mutation in the second exon of this gene has been reported to be associated with maximum variations in the kernel length. We have developed a simple PCR-based marker system named DRR-GL which targets the functional nucleotide polymorphism at GS3. This marker system has the advantages that it is easy to use, saves time and cost, and is amenable for large-scale marker-assisted selection for the trait of kernel length. Validation of this marker in a segregating population and 152 rice varieties, which includes 30 elite basmati varieties, reveals its effective co-segregation and association with the traits of kernel length as well as kernel elongation after cooking. We recommend utilization of this simple, low-cost marker system in breeding programs targeted at improvement of key rice grain quality traits, kernel length and kernel elongation.
Molecular Breeding
Depending on the position on the panicle, grain filling differs for spikelets on primary branches... more Depending on the position on the panicle, grain filling differs for spikelets on primary branches and secondary branches of upper and lower portions in rice (Oryza sativa L.). With the objective of identifying candidate genes associated with the grain-filling process across the panicle, a candidate gene (CG)-based mapping approach was attempted in an F 2 population derived from the cross between Rasi, a rice variety known for its good grain filling, and IC114927, a local landrace. The grain filling (%) was characterized for 444 F 2 individual plants across the panicle. Within the F 2 population, thirteen plants were identified with >90% grain filling across the panicle. For 120 markers designed from 1 kb upstream and within the candidate genes reported to be involved in translocation process of sugars, 18.3% polymorphism was observed. Using a selective genotyping approach comprising 24 individuals from eight groups based on their filling and position, two associated CG markers were identified. Further analysis of the mapping population showed a CG marker based on the sucrose phosphate synthase gene on chromosome 2 to be significantly associated with filling of grains on primary branches of the upper half of the panicle and another CG marker based on the transporter gene on chromosome 11 to be associated with filling of grains on primary branches of the lower half; secondary branches of the upper half and lower half of the panicle. Both the positive alleles were contributed by the variety Rasi. With a candidate gene-based marker mapping approach, two genes involved in sugar translocation and transport were identified to be directly associated with grain filling in rice.
During the past five decades, a large number of tobacco varieties have been developed for differe... more During the past five decades, a large number of tobacco varieties have been developed for different end uses in India through pure line selection from local land-races, mutation breeding and hybridization involving local selections and exotic introductions followed by pedi- gree selection. No systematic effort has been made to understand the existing diversity pattern in these va- rieties, which is
Euphytica, 2009
SSR markers targeting (GATA) n motifs are known to be highly polymorphic and useful in many organ... more SSR markers targeting (GATA) n motifs are known to be highly polymorphic and useful in many organisms. (GATA) n motif specific SSR markers covering the whole rice genome are not available. The present study was carried out with an objective to identify class I rice microsatellites in the rice genome with (GATA) n motifs, in-silico, and validate their potential as molecular markers. A total of 243 such motifs were identified; 65 of these were present in the genic region, 59 were in the upstream region and the remaining motifs were found in the intergenic regions. Many of the (GATA) n motifs were found within and/or upstream of genes associated with biotic or abiotic stress tolerance. A total of 230 PCRbased markers targeting all the class I (GATA) n microsatellites were developed and 35 of these markers spread across the rice genome were validated in a set of 24 representative rice varieties belonging to five distinct cultivar groups. All the markers were polymorphic, with average polymorphism information content (PIC) value of 0.61, and the rice cultivars could be uniquely distinguished into different cultivar groups based on marker analysis. These informative markers targeting (GATA) n motifs representing a new set of markers in rice will be highly useful for genetic studies and marker-assisted selection.
Proceedings of 2nd …, 2006
Molecular …, 2009
Fragrance development in rice has been reported due to a 8-bp deletion in the exon 7 of badh2 gen... more Fragrance development in rice has been reported due to a 8-bp deletion in the exon 7 of badh2 gene located on Chromosome 8S. Multiplex markers targeting the functional InDel polymorphism was earlier reported for genotyping fragrance trait, but the marker was observed to be inconsistent and difficult to use. We have developed a simple, co-dominant, functional marker for fragrance trait, which can be resolved in an agarose gel and validated in Basmati and non-Basmati aromatic rice varieties and in a mapping population segregated for fragrance trait. The marker targets the InDel polymorphism in badh2 gene and amplifies 95 and 103 bp fragments in fragrant and non-fragrant genotypes, respectively. The newly developed marker was highly efficient in discriminating all fragrant and non-fragrant genotypes and showed perfect co-segregation with the trait of fragrance in the mapping population. We recommend the use of this simple, low-cost marker in routine genotyping for fragrance trait in large scale breeding materials and germplasm.
Biotechnology Advances, 2010
Enormous sequence information is available in public databases as a result of sequencing of diver... more Enormous sequence information is available in public databases as a result of sequencing of diverse crop genomes. It is important to use this genomic information for the identification and isolation of novel and superior alleles of agronomically important genes from crop gene pools to suitably deploy for the development of improved cultivars. Allele mining is a promising approach to dissect naturally occurring allelic variation at candidate genes controlling key agronomic traits which has potential applications in crop improvement programs. It helps in tracing the evolution of alleles, identification of new haplotypes and development of allele-specific markers for use in marker-assisted selection. Realizing the immense potential of allele mining, concerted allele mining efforts are underway in many international crop research institutes. This review examines the concepts, approaches and applications of allele mining along with the challenges associated while emphasizing the need for more refined ‘mining’ strategies for accelerating the process of allele discovery and its utilization in molecular breeding.
Molecular Genetics and Genomics, 2005
In order to understand the molecular mechanisms involved in the gene-for-gene type of pathogen re... more In order to understand the molecular mechanisms involved in the gene-for-gene type of pathogen resistance, high-resolution genetic and physical mapping of resistance loci is required to facilitate map-based cloning of resistance genes. Here, we report the molecular mapping and cloning of a dominant gene (Pi-k h ) present in the rice line Tetep, which is associated with resistance to rice blast disease caused by Magnaporthe grisea. This gene is effective against M. grisea populations prevalent in the Northwestern Himalayan region of India. Using 178 sequence tagged microsatellite, sequence-tagged site, expressed sequence tag and simple sequence repeat (SSR) markers to genotype a population of 208 F2 individuals, we mapped the Pi-k h gene between two SSR markers (TRS26 and TRS33) which are 0.7 and 0.5 cM away, respectively, and can be used in marker-assisted-selection for blast-resistant rice cultivars. We used the markers to identify the homologous region in the genomic sequence of Oryza sativa cv. Nipponbare, and a physical map consisting of two overlapping bacterial artificial chromosome and P1 artificial chromosome clones was assembled, spanning a region of 143,537 bp on the long arm of chromosome 11. Using bioinformatic analyses, we then identified a candidate blast-resistance gene in the region, and cloned the homologous sequence from Tetep. The putative Pi-k h gene cloned from Tetep is 1.5 kbp long with a single ORF, and belongs to the nucleotide binding site-leucine rich repeat class of disease resistance genes. Structural and expression analysis of the Pi-k h gene revealed that its expression is pathogen inducible.