Construction of genetic linkage map and QTL analysis of sinksize traits in pearl millet (Pennisetum glaucum)

A linkage map, primarily based on SSCP-SNP markers, was constructed using 188 F2:3 (F2-derived F3) mapping population progenies derived from a cross between two pearl millet inbred lines having diverse pedigrees. The parents had large differences for two sink size traits (grain size and panicle diameter), and also differed for panicle length. The skeleton linkage map covered 1019 cM and it comprised of 44 loci (detected with 24 SSCP-SNP, 10 genomic SSR, 6 EST-SSR and 4 STS primer pairs) distributed across the seven linkage groups. Average adjacent-marker intervals ranged from 14 cM on LG1 to 38 cM on LG6, with an overall mean of 23 cM. Using the F2 linkage map and phenotypic data collected from the F2 and F2:3 generations of the mapping population, a total of 18 putative QTLs were detected for the three sink-size components. Eight QTLs explained 42.7% of observed phenotypic variation for panicle length, with individual QTLs explaining 6.1 to 18.2% using the F2:3 data set. For panicle diameter, 5 QTLs explained 45.8% of observed phenotypic variation with individual QTLs accounting for 6.3 to 30.2%. Similarly for grain size, 5 QTLs explained 29.6% of phenotypic variation with individual QTLs accounting for 6.1 to 8.9%. Genomic regions associated with panicle length, panicle diameter and grain size co-mapped on LG6 between Xpsms88 and Xpsms2270, indicating the existence of a gene or gene cluster with major effects involved in the control of significant proportions of the phenotypic variation for all three sink-size traits. The QTLs for panicle length on LG2 and LG6 (LOD>3 in both F2 and F2:3 data sets), for panicle diameter on LG2 and LG3 (LOD>14 in the F2:3 data set) and for grain size on LG3 and LG6 (LOD>3 in both F2 and F2:3 data sets) were identified as promising candidates for validation prior to possible application in marker-assisted breeding

Bioefficacy of certain chemical and biofungicides against Hypoxylon spp. causing wood rot disease in tea

Wood rot disease caused by Hypoxylon serpens is the most widespread and serious stem disease in tea. Among the 350 bacterial and 35 fungal biocontrol isolates collected from several tea growing regions of southern India, three bacterial isolates produced higher antagonistic potential against this fungal pathogen. Two of the efficient strains were identified as Bacillus sp. (HBCWR-3 and WR46-2) and third one was Pseudomonas sp. (WR5-4). In case of fungal biocontrol agents, the type culture Trichoderma viride procured from Microbial Type Culture Collection (MTCC) performed better in controlling the pathogen over T. harzianum. Five systemic fungicides, hexaconazole, carbendazim, tebuconazole, tridemorph, benomyl and a contact fungicide, copper oxychloride were evaluated for studying their bioefficacy against wood rot pathogen. In this study, benomyl 50% WP or copper oxychloride at the lowest concentration (0.01%) completely inhibited the growth of the fungus in vitro. Moreover, bioefficacy of certain plant aqueous extracts of Azadirachta indica, acetone extracts of Pongamia pinnata, Cinnamom, Artemisia nilagirica, Lantana camera, Ageratum conyzoides and a bryophyte, Heteroscyphus argutus were also studied against H. serpens. Among them, A. nilagirica followed by H. argutus and A. indica were effective in controlling the wood rot pathogen. In the case of liquid biofungicides tested, ‘Expel’ controlled the tea pathogen efficiently. The present study revealed that, chemical fungicide (Benomyl or copper oxychloride at 0.01%), botanical extracts at 10% (A. nilagirica, H. argutus, Azadirachta and ‘Expel’) and biocontrol agents (Bacillus sp., Pseudomonas sp. and T. viride) were effective in controlling wood rot pathogen under in vitro condition

Bioefficacy of certain chemical and biofungicides against Hypoxylon spp. causing wood rot disease in tea

Wood rot disease caused by Hypoxylon serpens is the most widespread and serious stem disease in tea. Among the 350 bacterial and 35 fungal biocontrol isolates collected from several tea growing regions of southern India, three bacterial isolates produced higher antagonistic potential against this fungal pathogen. Two of the efficient strains were identified as Bacillus sp. (HBCWR-3 and WR46-2) and third one was Pseudomonas sp. (WR5-4). In case of fungal biocontrol agents, the type culture Trichoderma viride procured from Microbial Type Culture Collection (MTCC) performed better in controlling the pathogen over T. harzianum. Five systemic fungicides, hexaconazole, carbendazim, tebuconazole, tridemorph, benomyl and a contact fungicide, copper oxychloride were evaluated for studying their bioefficacy against wood rot pathogen. In this study, benomyl 50% WP or copper oxychloride at the lowest concentration (0.01%) completely inhibited the growth of the fungus in vitro. Moreover, bioefficacy of certain plant aqueous extracts of Azadirachta indica, acetone extracts of Pongamia pinnata, Cinnamom, Artemisia nilagirica, Lantana camera, Ageratum conyzoides and a bryophyte, Heteroscyphus argutus were also studied against H. serpens. Among them, A. nilagirica followed by H. argutus and A. indica were effective in controlling the wood rot pathogen. In the case of liquid biofungicides tested, ‘Expel’ controlled the tea pathogen efficiently. The present study revealed that, chemical fungicide (Benomyl or copper oxychloride at 0.01%), botanical extracts at 10% (A. nilagirica, H. argutus, Azadirachta and ‘Expel’) and biocontrol agents (Bacillus sp., Pseudomonas sp. and T. viride) were effective in controlling wood rot pathogen under in vitro condition

Bioefficacy of efficient entomopathogenic fungus against branch canker pathogen (Macrophoma theicola) in tea plantations of southern India

Three branch canker pathogens, viz. NBCHE-6, UPA-61 and VPM were isolated from different tea growing districts of south India and four entomopathogenic fungus, viz. Beauveria bassiana, Paecilomyces lilacinus, Lecannicillium lecannii and Paecilomyces fumosoroseus were procured from the microbial type culture collection and gene bank (MTCC), Chandigarh. In vitro studies revealed that Beuveria bassiana showed highest antagonistic effect against NBCHE-6 (64.22) followed by Paecilomyces fumosoroseus against UPA-61(56.66). Paecilomyces lilacinus significantly controlled VPM (54.66), while Lecannicillium lecannii showed insignificantly control against VPM (47.33). While Beuveria bassiana and Paecilomyces lilacinus coiled around and shrink branch canker pathogen, Lecannicillium lecannii breaks into branch canker hyphae and Paecilomyces fumosoroseus produces more spore to kill branch canker. In culture filtrate studies, Paecilomyces fumosoroseus and Paecilomyces lilacinus showed maximum control of VPM (68.44) and UPA-61 (65.59). Beauveria bassiana also showed significant control of two isolates VPM and UPA-61 (54.44). Lecanniicillium lecannii showed least control of VPM (30.44). This study concludes that entomopathogens can significantly control branch canker pathogen (Macrophoma theicola)

Phenotypic and molecular diversity-based prediction of heterosis in pearl millet ( Pennisetum glaucum L. (R.) Br.)

Genetic distances between hybrid parents based on phenotypic traits and molecular markers were investigated to assess their relationship with heterosis for grain and stover yield and other traits in pearl millet (Pennisetum glaucum [L.] R. Br.). Fifty-one hybrids developed using 101 hybrid parents (B and R lines) and showing a wide range of genetic distance between their parents based on eight phenotypic traits and 28–38 SSRs were evaluated in two sets for two seasons. The correlation between Euclidean distance (phenotypic distance, ED) and simple matching distance (molecular distance, SM) for parents of both sets was low but positive and significant (r = 0.2, P < 0.001).The correlation of ED in parents with better-parent heterosis for grain yield was similar in both sets (r = 0.38, P < 0.05). SM was not correlated with heterosis for grain yield in either set of hybrids. The results showed that phenotypic distance could be a better predictor of heterosis than molecular distance. The correlation between phenotypic distance and heterosis was not strong enough to permit the use of phenotypic diversity among parents as a major selection criterion for selection of parental lines displaying high levels of heterosis for grain and stover yield in pearl millet

Genetic Diversity in Maintainer and Restorer Lines of Pearl Millet

Molecular markers facilitate rapid and environment-neutral characterization of the pattern of genetic diversity. The International Crops Research Institute for the Semi-Arid Tropics (ICRISAT) at Patancheru, India, has developed a large number and diverse range of maintainer lines (B-lines) and restorer lines (R-lines) of pearl millet [Pennisetum glaucum (L.) R. Br.] in last three decades. In the present study, 213 lines (98 B-lines and 115 R-lines) were genotyped using high throughput assay and 38 polymorphic simple sequence repeat (SSR) markers, which detected 308 alleles averaging 8.1 alleles per locus. Restorer lines were genetically more diverse than B-lines, as higher average gene diversity was detected among R-lines (0.62) than among B-lines (0.50). A neighbor-joining tree based on simple matching dissimilarity distance matrix clearly differentiated B-lines from R-lines into two clusters, with further subdivision of B-lines into four subclusters and R-lines into five subclusters. Seven B-lines clustered with R-lines, and only two R-lines clustered with B-lines. Few unique alleles (1 to 3) were detected in 14 B- and 30 R-lines, which could be used to distinguish them from each other as well as from the remaining lines used in this study. The SSR alleles of each of the parental lines along with distinctness, uniformity, and stability (DUS) characterization may enable ICRISAT to protect these lines from any kind of infringement by the multiple users who have received these lines for hybrid development

Patterns of Molecular Diversity in Current and Previously Developed Hybrid Parents of Pearl Millet [Pennisetum glaucum (L.) R. Br.]

ICRISAT’s pearl millet (Pennisetum glaucum (L.) R. Br.) breeding program at Patancheru, India, has developed genetically diverse hybrid parents since 1980s. The present study investigated genetic diversity pattern between two groups of parents in this program, bred till 2004 and developed during 2004-2010. Combined analysis of 379 hybrid parents (current 166 parents and 213 previously developed hybrid parents) carried out using a set of highly polymorphic 28 SSRs detected 12.7 alleles per locus. An average of 8.5 and 8.7 SSR alleles per locus were found in previously developed and current parents, respectively, indicating marginal improvement in the levels of genetic diversity of hybrid parents in this program. Distance matrix differentiated these current and previously developed hybrid parents into 2 separate clusters, indicating infusion of new genetic variability over time as reflected by development of more genotype-specific alleles. Also, the seed and restorer parents were found clearly separated from each other in both the sets with few crossovers, indicating existence of two diverse and broad-based pools in hybrid parents of pearl millet. Restorer parents (R-lines) were found more diverse than seed parents (B-lines), as higher average gene diversity was detected among R-lines (0.70) than B-lines (0.56), though variation between B- and R-lines was found reduced in newly developed lines to 9.22% from 16.98% in previously developed lines. Results suggested that newly developed lines were as much divergent when compared with previously developed lines, indicating that current ICRISAT pearl millet breeding program was moving towards development of diverse new hybrid parental lines. The study suggested use of trait-specific donors in B- and R-lines separately to maintain sufficient genetic distance between seed and restorer breeding lines. It was pointed out to cross parents having higher genetic distance within the seed (B-lines) and restorer (R-lines) breeding programs to derive diverse and productive hybrid parental lines in future

Identification of genes controlling compatible and incompatible reactions of pearl millet (Pennisetum glaucum) against blast (Magnaporthe grisea) pathogen through RNA-Seq

Blast [Magnaporthe grisea (Herbert) Barr] is an economically important disease in Asian pearl millet production ecologies. The recurrent occurrence of blast in the past one decade has caused enormous strain on grain and forage production. Identification of resistance genes is an important step to develop durable varieties. The present study is the first attempt to use RNA-Seq to investigate the transcript dynamics in a pearl millet inbred ICMB 93333, which had a unique differential reaction to two isolates—Pg 45 (avirulent) and Pg 174 (virulent) of M. grisea. The inbred was inoculated by both isolates and samples taken at six different time intervals for genome-wide RNA-Seq experiment. The transcriptome results revealed the differential expression of more than 2,300 genes. The time-specific comparison showed activation or repression of specific genes in various pathways. Genes and transcriptions factors related to pathogenesis-related proteins, reactive oxygen species generating and its scavenging genes, cell wall defense, primary and secondary metabolic pathways, and signaling pathways were identified by comparing the host-plant compatible and incompatible interactions. The genes identified from this experiment could be useful to understand the host-plant resistance and design novel strategies to manage blast disease in pearl millet

Morphological, Chemical and In Vitro Traits for Prediction of Stover Quality in Pearl Millet for Use in Multidimensional Crop Improvement

The paper reports on the variations in organic matter digestibility (OMD), organic matter intake (OMI), digestible organic matter intake (DOMI) and nitrogen (N) balances of 40 pearl millet stover fed to sheep ad libitum and investigated relationships between these in vivo variables and morphological, chemical and in vitro stover quality traits. Highly significant differences (P<0.0001) were observed for all in vivo variables. Plant height and stem diameter were consistently significantly (at least P=0.0002) inversely related to OMD, OMI, DOMI and N balance. Except for OMD (P=0.16) stover nitrogen content was significantly (at least P=0.0005) positively related to OMI, DOMI and N balance. The fiber constituents neutral (NDF) and acid (ADF) detergent fiber and acid detergent lignin (ADL) were consistently inversely related to OMD, OMI, DOMI and N balance. (at least P=0.0005). Stover in vitro digestibility, metabolizable energy content and extent and rate of in vitro gas production were highly positively (at least P<0.0009) related to the in vivo variables while lag and half time of in vitro gas production were negatively associated (at least P=0.0004) with the in vivo variables. Combined morphological, chemical and in vitro variables in stepwise multiple regressions accounted for 70 to 84%% of the variations observed in the in vivo variables. Application of stringent cross validation procedures reduced the variation in OMD, OMI, DOMI and N balance accounted for by the combined morphological, chemical and in vitro variables mostly moderately to 71, 49, 79 and 76%, respectively. Relatively simple traits can predict stover quality in breeding programs for dual-purpose pearl millet cultivar

Construction of Genetic Linkage Map and QTL Analysis of Sink-Size Traits in Pearl Millet (Pennisetum glaucum)

A linkage map, primarily based on SSCP-SNP markers, was constructed using 188 F2:3 mapping population progenies produced from a cross between two pearl millet inbred lines having diverse parentage. The skeleton linkage map covered 1019 cM and it comprised of 44 markers distributed across the seven linkage groups. Average adjacent-marker intervals ranged from 14 cM on LG1 to 38 cM on LG6, with an overall mean of 23 cM. Using the F2 linkage map and phenotypic data from the F2 and F2:3 generations of the mapping population, a total of 18 putative QTLs were detected for the three sink-size components. Eight QTLs explained 42.7% of observed phenotypic variation for panicle length using the F2:3 data set. For panicle diameter, 5 QTLs explained 45.8% of observed phenotypic variation. Similarly for grain size, 5 QTLs explained 29.6% of phenotypic variation. Genomic regions associated with panicle length, panicle diameter, and grain size were comapped on LG6 between Xpsms88 and Xpsms2270, indicating the existence of a gene or gene cluster. The QTLs for panicle length on LG2 and LG6 ( in both F2 and F2:3 data sets), for panicle diameter on LG2 and LG3 ( in the F2:3 data set), and for grain size on LG3 and LG6 ( in both F2 and F2:3 data sets) were identified as promising candidates for validation prior to possible application in marker-assisted breeding