Firoz Hossain

Genome-wide identification and expression pattern of drought-responsive members of the NAC family in maize

NAC proteins are plant-specific transcription factors (TFs). Although they play a pivotal role in regulating distinct biological processes, TFs in maize are yet to be investigated comprehensively. Within the maize genome, we identified 152 putative NAC domain-encoding genes (ZmNACs), including eight membrane-bound members, by systematic sequence analysis and physically mapped them onto ten chromosomes of maize. In silico analysis of the ZmNACs and comparison with similar genes in other plants such as Arabidopsis, rice, and soybean, revealed a similar NAC sequence architecture. Phylogenetically, the ZmNACs were arranged into six distinct subgroups (I–VI) possessing conserved motifs. Phylogenetic analysis using stress-related NAC TFs from Arabidopsis, rice, and soybean as seeding sequences identified 24 of the 152 ZmNACs (all from Group II) as putative stress-responsive genes, including one dehydration-responsive ZmSNAC1 gene reported earlier. One drought-tolerant genotype (HKI577) and one susceptible genotype (PC13T-3) were used for studying the expression pattern of the NAC genes during drought stress. qRT-PCR based expression profiles of 11 genes predicted to be related to stress confirmed strong differential gene expression during drought stress. Phylogenetic analyses revealed that ZmNAC18, ZmNAC51, ZmNAC145, and ZmNAC72, which were up-regulated in the tolerant genotype and down-regulated in the susceptible genotype, belonged to the same group to which also belong other drought-responsive genes, namely SNAC1, OsNAC6, ANAC019, and ANAC055, which act as a transcriptional activator and are strongly induced under stress from various abiotic sources. Differentially expressed ZmNAC genes, alone or in combination with each other or with other type(s) of TFs, may control the general cellular machinery and regulate stress-responsive downstream genes. Alternatively, they may serve as a platform to regulate a broad set of genes, which are subsequently fine-tuned by specific regulators. This genome-wide identification and expression profiling opens new avenues for systematic functional analysis of new members of the NAC gene family, which may be exploited in developing lines that are better adapted to drought.

Development of β-carotene rich maize hybrids through marker-assisted introgression of β-carotene hydroxylase allele.

Development of vitamin A-rich cereals can help in alleviating the widespread problem of vitamin A deficiency. We report here significant enhancement of kernel β-carotene in elite maize genotypes through accelerated marker-assisted backcross breeding. A favourable allele (543 bp) of the β-carotene hydroxylase (crtRB1) gene was introgressed in the seven elite inbred parents, which were low (1.4 µg/g) in kernel β-carotene, by using a crtRB1-specific DNA marker for foreground selection. About 90% of the recurrent parent genome was recovered in the selected progenies within two backcross generations. Concentration of β-carotene among the crtRB1-introgressed inbreds varied from 8.6 to 17.5 µg/g - a maximum increase up to 12.6-fold over recurrent parent. The reconstituted hybrids developed from improved parental inbreds also showed enhanced kernel β-carotene as high as 21.7 µg/g, compared to 2.6 µg/g in the original hybrid. The reconstituted hybrids evaluated at two locations possessed similar grain yield to that of original hybrids. These β-carotene enriched high yielding hybrids can be effectively utilized in the maize biofortification programs across the globe.

Genome-Wide Expression of Transcriptomes and Their Co-Expression Pattern in Subtropical Maize (Zea mays L.) under Waterlogging Stress

Waterlogging causes extensive damage to maize crops in tropical and subtropical regions. The identification of tolerance genes and their interactions at the molecular level will be helpful to engineer tolerant genotypes. A whole-genome transcriptome assay revealed the specific role of genes in response to waterlogging stress in susceptible and tolerant genotypes. Genes involved in the synthesis of ethylene and auxin, cell wall metabolism, activation of G-proteins and formation of aerenchyma and adventitious roots, were upregulated in the tolerant genotype. Many transcription factors, particularly ERFs, MYB, HSPs, MAPK, and LOB-domain protein were involved in regulation of these traits. Genes responsible for scavenging of ROS generated under stress were expressed along with those involved in carbohydrate metabolism. The physical locations of 21 genes expressed in the tolerant genotype were found to correspond with the marker intervals of known QTLs responsible for development of adaptive traits. Among the candidate genes, most showed synteny with genes of sorghum and foxtail millet. Co-expression analysis of 528 microarray samples including 16 samples from the present study generated seven functional modules each in the two genotypes, with differing characteristics. In the tolerant genotype, stress genes were co-expressed along with peroxidase and fermentation pathway genes.

Functional mechanisms of drought tolerance in subtropical maize (Zea mays L.) identified using genome-wide association mapping

BackgroundEarlier studies were focused on the genetics of temperate and tropical maize under drought. We identified genetic loci and their association with functional mechanisms in 240 accessions of subtropical maize using a high-density marker set under water stress.ResultsOut of 61 significant SNPs (11 were false-discovery-rate-corrected associations), identified across agronomic traits, models, and locations by subjecting the accessions to water stress at flowering stage, 48% were associated with drought-tolerant genes. Maize gene models revealed that SNPs mapped for agronomic traits were in fact associated with number of functional traits as follows: stomatal closure, 28; flowering, 15; root development, 5; detoxification, 4; and reduced water potential, 2. Interactions of these SNPS through the functional traits could lead to drought tolerance. The SNPs associated with ABA-dependent signalling pathways played a major role in the plant’s response to stress by regulating a series of functions including flowering, root development, auxin metabolism, guard cell functions, and scavenging reactive oxygen species (ROS). ABA signalling genes regulate flowering through epigenetic changes in stress-responsive genes. ROS generated by ABA signalling are reduced by the interplay between ethylene, ABA, and detoxification signalling transductions. Integration of ABA-signalling genes with auxin-inducible genes regulates root development which in turn, maintains the water balance by regulating electrochemical gradient in plant.ConclusionsSeveral genes are directly or indirectly involved in the functioning of agronomic traits related to water stress. Genes involved in these crucial biological functions interacted significantly in order to maintain the primary as well as exclusive functions related to coping with water stress. SNPs associated with drought-tolerant genes involved in strategic biological functions will be useful to understand the mechanisms of drought tolerance in subtropical maize.

Cyanobacterial inoculation elicits plant defense response and enhanced Zn mobilization in maize hybrids

Abstract The present investigation evaluated the effect of inoculating different cyanobacterial formulations on a set of hybrids of maize, in terms of plant defense enzyme activity, soil health parameters, Zn concentration, and yields. Microbial inoculation showed significant effects on accumulation of Zn in flag leaf, with A4 (Anabaena–Azotobacter biofilm) recording the highest values. Analysis of variance (ANOVA) indicated that both the hybrids and cyanobacterial treatments brought about significant variation in terms of glomalin-related soil proteins and polysaccharides in soil and the activity of defense enzymes in roots and shoots of the plants. Cyanobacterial inoculants—A4 (Anabaena–Azotobacter biofilm) and A1 (Anabaena sp.–Providencia sp., CW1 + PW5) enhanced the activity of peroxidase, PAL and PPO in roots, which also showed a positive correlation with Zn concentration in the flag leaf. Grain yield ranged from 7.0 to 7.29 t/ha among the different inoculants. Comparative analyses of treatments showed that A3 (Anabaena–Trichoderma-biofilmed formulation) and hybrid B8 (Bio-9681) were superior in terms of parameters investigated. This represents the first report on the genotypic responses of maize hybrids to cyanobacteria-based inoculants. Future research should focus on dissecting the role of root exudates and cyanobacteria-mediated Zn mobilization pathway in maize.

Identification of kernel iron- and zinc-rich maize inbreds and analysis of genetic diversity using microsatellite markers

Development of micronutrient enriched staple foods is an important breeding goal in view of the extensive problem of ‘hidden hunger’ caused by micronutrient malnutrition. In the present study, kernel iron (Fe) and zinc (Zn) concentrations were evaluated in a set of 31 diverse maize inbred lines in three trials at two locations – Delhi (Kharif 2007 & 2008) and Hyderabad (Rabi 2007–08). The ranges of kernel Fe and Zn concentrations were 13.95–39.31 mg/kg and 21.85–40.91 mg/kg, respectively, across the three environments. Pooled analysis revealed significant genotype × environment (G × E) interaction in the expression of both the micronutrient traits, although kernel Fe was found to be more sensitive to G × E as compared to kernel Zn. Seven inbred lines, viz., BAJIM-06-03, DQPM-6, CM212, BAJIM-06-12, DQPM-7, DQPM-2 and CM129, were found to be the most stable and promising inbred lines for kernel Zn concentration, while for kernel Fe concentration, no promising and stable genotypes could be identified. Analysis of molecular diversity in 24 selected inbred lines with phenotypic contrast for the two kernel micronutrient traits, using 50 SSR markers covering the maize genome, revealed high levels of polymorphisms (214 SSR alleles; mean PIC value = 0.62). The phenotypically contrasting and genetically diverse maize inbred lines identified in this study could be potentially utilized in further studies on QTL analysis of kernel micronutrient traits in maize, and the stable and most promising kernel micronutrient-rich maize genotypes provide a good foundation for developing micronutrient-enriched maize varieties suitable for the Indian context.

Evaluation of quality protein maize genotypes for resistance to stored grain weevil Sitophilus oryzae (Coleoptera: Curculionidae)

The stored grain weevil, Sitophilus oryzae (Linnaeus), causes significant losses to maize grains in tropical countries. Despite the nutritional superiority of the quality protein maize (QPM), an important concern is the possible vulnerability of QPM genotypes to stored grain weevil infestation. In the present study the responses of 24 QPM inbred lines, along with several non-QPM inbred lines and hybrids, were evaluated against S. oryzae under laboratory conditions. Two attributes, namely (i) % weight loss in the grains after a fixed period of incubation with the insect and (ii) the number of progeny insects (insect multiplication) generated from six pairs of adult insects, were considered. A cumulative resistance index was computed for each genotype giving equal weight to both attributes. The study revealed DMRQPM-60 and CML167 as the most resistant entries. Shaktiman-1, the released QPM hybrid in India, showed moderate resistance against the grain weevil. However, most of the elite QPM inbred lines as well as some non-QPM inbred lines and hybrids were found to be highly susceptible to the weevil infestation. To explore the possible relationship between enhanced protein quality and varying degree of kernel vitreousness and vulnerability to the grain weevil infestation, both the QPM and the non-QPM genotypes were analysed. The correlations between kernel quality traits and kernel texture with % kernel weight loss due to weevil infestation and insect multiplication were found to be non-significant, indicating possible influence of other factors, including pericarp thickness and endosperm composition, in determining the susceptibility of the QPM genotypes to S. oryzae. The study indicates that QPM genotypes are not necessarily susceptible to the stored grain weevil and identifies promising QPM genotypes with resistance to S. oryzae.

Enhancement of plant growth and yields in Chickpea (Cicer arietinum L.) through novel cyanobacterial and biofilmed inoculants.

The use of Rhizobium inoculants in chickpea is well established; however, meagre efforts have been directed towards the use of other microbial supplements for improving nutrient uptake and yields. A set of novel cyanobacterial and biofilmed inoculants were evaluated in chickpea under field conditions. A significant two-fold enhancement in leghaemoglobin content of nodules and plant biomass was recorded with Anabaena laxa treatment. The inoculants - Anabaena laxa and Anabaena - Rhizobium biofilmed formulation proved to be the top-ranking treatments. Soil chlorophyll, nitrogen-fixation and available N possessed high positive direct effects on grain yield through positive - correlations and - high direct effects and also had high positive indirect effects through other component traits. The cumulative effect of improved plant growth and nutrient uptake exhibited a positive correlation with microbiological activity, especially nitrogen fixation, soil chlorophyll and soil available nitrogen. This may account for the significantly higher yield parameters in the A. laxa treatment, which recorded 50% higher grain yield (1724kgha(-1)) as compared to control (847kgha(-1)).

Molecular characterization and assessment of genetic diversity of inbred lines showing variability for drought tolerance in maize

A set of 24 genotypes bred at different centres in India as well as in CIMMYT showing variability for drought tolerance were selected for molecular and morpho-physiological characterization. A set of 35 SSR markers, having genome-wide coverage, was chosen for genotyping the inbreds. These markers generated a total of 111 polymorphic alleles with an average of 3.17 alleles per locus. The minimum and maximum PIC value was 0.27 and 0.77 with a mean of 0.5. A total of 13 unique alleles were found in the 24 inbred lines. The coefficient of genetic dissimilarity ranged from 0.192 to 0.803. NJ-based tree suggested the presence of three major clusters of which, two of them had subgroups. Phenotyping of inbreds by morpho-physiological traits revealed that there was a positive relationship among root length, chlorophyll content, relative water content while anthesis-silking interval was negative relationship with all these traits. Genotyping data complemented by morpho-physiological parameters were used to identify a number of pair-wise combinations for the development of mapping population segregating for drought tolerance and potential heterotic pairs for the development of drought tolerant hybrids.

Beneficial cyanobacteria and eubacteria synergistically enhance bioavailability of soil nutrients and yield of okra

Microorganisms in the rhizosphere mediate the cycling of nutrients, their enhanced mobilisation and facilitate their uptake, leading to increased root growth, biomass and yield of plants. We examined the promise of beneficial cyanobacteria and eubacteria as microbial inoculants, applied singly or in combination as consortia or biofilms, to improve growth and yields of okra. Interrelationships among the microbial activities and the micro/macro nutrient dynamics in soils and okra yield characteristics were assessed along with the changes in the soil microbiome. A significant effect of microbial inoculation on alkaline phosphatase activity was recorded both at the mid-crop and harvest stages. Microbial biomass carbon values were highest due to the Anabaena sp. - Providencia sp. (CR1 + PR3) application. The yield of okra ranged from 444.6–478.4 g−1 plant and a positive correlation (0.69) recorded between yield and root weight. The application of Azotobacter led to the highest root weight and yield. The concentration of Zn at mid-crop stage was 60–70% higher in the Azotobacter sp. and Calothrix sp. inoculated soils, as compared to uninoculated control. Iron concentration in soil was more than 2–3 folds higher than control at the mid-crop stage, especially due to the application of Anabaena-Azotobacter biofilm and Azotobacter sp. Both at the mid-crop and harvest stages, the PCR-DGGE profiles of eubacterial communities were similar among the uninoculated control, the Anabaena sp. - Providencia sp. (CW1 + PW5) and the Anabaena-Azotobacter biofilm treatments. Although the profiles of the Azotobacter, Calothrix and CR1 + PR3 treatments were identical at these stages of growth, the profile of CR1 + PR3 was clearly distinguishable. The performance of the inoculants, particularly Calothrix (T6) and consortium of Anabaena and Providencia (CR1 + PR3; T5), in terms of microbiological and nutrient data, along with generation of distinct PCR-DGGE profiles suggested their superiority and emphasized the utility of combining microbiological and molecular tools in the selection of effective microbial inoculants.