ABSTRACT
The human gut microbiota can biosynthesize essential micronutrients such as B-vitamins and is also known for its metabolic cooperative behaviour. The present study characterises such B-vitamin biosynthesizers, their biosynthetic pathways, explores their prevalence and abundance, examines how lifestyle or diet affects them in multiple Indian cohorts and compares it with the Chinese cohort. To achieve this, publicly available faecal metagenome data of healthy individuals from multiple Indian (two urban and three tribal populations) and a Chinese cohort were analysed. The distribution of prevalence and abundance of B-vitamin biosynthesizers showed similar profiles to that of the entire gut community of the Indian cohort, and there were 28 B-vitamin biosynthesizers that had modest or higher prevalence and abundance. The omnivorous diet affected only the prevalence of a few B-vitamin biosynthesizers; however, lifestyle and/or location affected both prevalence and abundance. A comparison with the Chinese cohort showed that fourteen B-vitamin biosynthesizers were significantly more prevalent and abundant in Chinese as compared with Indian samples (False Discovery Rate (FDR) <= 0·05). The metabolic potential of the entire gut community for B-vitamin production showed that within India, the tribal cohort has a higher abundance of B-vitamin biosynthesis pathways as compared with two urban cohorts namely, Bhopal and Kasargod, and comparison with the Chinese cohort revealed a higher abundance in the latter group. Potential metabolic cooperative behaviour of the Indian gut microbiome for biosynthesis of the B-vitamins showed multiple pairs of species showed theoretical complementarity for complete biosynthetic pathways genes of thiamine, riboflavin, niacin and pantothenate.
Subject(s)
Gastrointestinal Microbiome , Vitamin B Complex , Humans , Gastrointestinal Microbiome/genetics , Thiamine , Riboflavin/analysis , ChinaABSTRACT
KEY MESSAGE: Genome-wide structural variants we identified and new NOR-linked markers we developed would be useful for future genome-wide association studies (GWAS), and for new gene/trait mapping purposes. Bioinformatic alignment of the assembled genomes of Col-0 and Sha ecotypes of Arabidopsis thaliana revealed ~ 13,000 genome-wide structural variants involving simple insertions or deletions and repeat contractions or expansions. Using some of these structural variants, we developed new, rapid, and low-cost PCR-based molecular markers that are genetically linked to the nucleolus organizer regions (NORs). A. thaliana has two NORs, one each on chromosome 2 (NOR2) and chromosome 4 (NOR4). Both NORs are ~ 4 Mb each, and hundreds of 45S ribosomal RNA (rRNA) genes are tandemly arrayed at these loci. Using previously characterized recombinant inbred lines (RILs) derived from Sha x Col-0 crosses, we validated the utility of the newly developed NOR-linked markers in genetically mapping rRNA genes and the associated telomeres to either NOR2 or NOR4. Lastly, we sequenced Sha genome using Oxford Nanopore Technology (ONT) and used the data to obtain sequences of NOR-telomere junctions, and with the help of RILs, we mapped them as new genetic markers to their respective NORs (NOR2-TEL2N and NOR4-TEL4N). The structural variants obtained from this study would serve as valuable data for genome-wide association studies (GWAS), and to rapidly design more genome-wide genetic (molecular) markers for new gene/trait mapping purposes.
Subject(s)
Arabidopsis , Arabidopsis/genetics , Genome-Wide Association Study , Chromosome Mapping , Base Sequence , TelomereABSTRACT
During amoebiasis, colonization of the gut by Entamoeba histolytica can lead to alterations of the host microbiota. In this study, we have compared the gut microbiota of patients of amoebiasis with healthy controls using 16S rRNA gene variable regions, (V1-V3) and (V3-V5), of the bacterial genome. From this 16S rRNA gene amplicon data, one paired-end and two single-end datasets were selected and compared by the number of OTUs obtained, sequence count, and diversity analysis. Our results showed that the V1-V3-paired-end dataset gave the maximum number of OTUs in comparison to the two single-end datasets studied. The amoebiasis samples showed a significant drop in richness in the alpha diversity measurements and lower intra group similarity compared to the healthy controls. Bacteria of genus Prevotella, Sutterella, and Collinsella were more abundant in healthy controls whereas Escherichia, Klebsiella, and Ruminococcus were more abundant in the E. histolytica-positive patients. All the healthy controls harbored bacteria belonging to Faecalibacterium, Prevotella, Ruminococcus, Subdoligranulum, and Escherichia genera while all the E. histolytica-positive patient samples contained genus Enterobacter. The compositional changes in the gut microbiome observed in our study indicated a higher prevalence of pathogenic bacteria along with a depletion of beneficial bacteria in E. histolytica-infected individuals when compared with healthy controls. These results underline the interplay between E. histolytica and the human gut microbiome, giving important inputs for future studies and treatments.
Subject(s)
Entamoebiasis , Gastrointestinal Microbiome , Humans , Gastrointestinal Microbiome/genetics , RNA, Ribosomal, 16S/genetics , Bacteria/genetics , Diarrhea , India , Feces/microbiologyABSTRACT
BACKGROUND: Intrapartum care is of paramount importance in overall respectful maternity care (RMC). Uterine fundal pressure maneuver (UFPM) is the most controversial obstetric maneuver considered obsolete in many countries. UFPM is associated with adverse life threatening maternal and fetal effects. The baseline prevalence of UFPM in our tertiary care teaching hospital with a heavy patient load was 78.4% which was quite high. Our aim was to reduce the prevalence of UFPM by 75% from baseline over a period of 10 months. METHODS: After root cause analysis, we formulated the departmental Standard Operating Procedures on safe vaginal delivery and birth practices and initiated the Labour Room Quality Improvement Initiative. We sensitized and created awareness among the resident doctors and nursing staff regarding the high prevalence of UFPM through dedicated lectures, intercommunication via WhatsApp groups and strict vigilance in the labor wards. The point of care quality improvement (QI) methodology was used. The primary outcome was decrease in the prevalence of UFPM. RESULTS: The prevalence of UFPM reduced from the baseline value of 78.4%-4% over the period of 7 months. Post intervention it gradually increased and sustained at 21.2% over a follow-up period of 3 months. CONCLUSION: QI methods can effectively and rapidly improve the acceptance and adherence to newer initiatives in a busy tertiary care health facility to reduce the prevalence of UFPM and eventually improve the overall RMC.
Subject(s)
Maternal Health Services , Quality Improvement , Pregnancy , Female , Humans , Prevalence , Developing Countries , Delivery, Obstetric/methods , Labor Stage, Second , Quality of Health CareABSTRACT
MAIN CONCLUSION: Comparative analysis of genome-wide miRNAs and their gene targets between cytoplasmic male sterile (CMS) and fertile lines of pigeonpea suggests a possible role of miRNA-regulated pathways in reproductive development. Exploitation of hybrid vigor using CMS technology has delivered nearly 50% yield gain in pigeonpea. Among various sterility-inducing cytoplasms (A1-A9) reported so far in pigeonpea, A2 and A4 are the two major sources that facilitate hybrid seed production. Recent evidence suggests involvement of micro RNA in vast array of biological processes including plant reproductive development. In pigeonpea, information about the miRNAs is insufficient. In view of this, we sequenced six small RNA libraries of CMS line UPAS 120A and isogenic fertile line UPAS 120B using Illumina technology. Results revealed 316 miRNAs including 248 known and 68 novel types. A total of 637 gene targets were predicted for known miRNAs, while 324 genes were associated with novel miRNAs. Degradome analysis revealed 77 gene targets of predicted miRNAs, which included a variety of transcription factors playing key roles in plant reproduction such as F-box family proteins, apetala 2, auxin response factors, ethylene-responsive factors, homeodomain-leucine zipper proteins etc. Differential expression of both known and novel miRNAs implied roles for both conserved as well as species-specific players. We also obtained several miRNA families such as miR156, miR159, miR167 that are known to influence crucial aspects of plant fertility. Gene ontology and pathway level analyses of the target genes showed their possible implications for crucial events during male reproductive development such as tapetal degeneration, pollen wall formation, retrograde signaling etc. To the best of our knowledge, present study is first to combine deep sequencing of small RNA and degradome for elucidating the role of miRNAs in flower and male reproductive development in pigeonpea.
Subject(s)
Cajanus/genetics , MicroRNAs , Plant Infertility/genetics , RNA, Plant/genetics , Cajanus/physiology , Cytoplasm , Gene Expression Profiling , Gene Expression Regulation, Plant , High-Throughput Nucleotide Sequencing , MicroRNAs/geneticsABSTRACT
In C4 species, ß-carbonic anhydrase (CA), localized to the cytosol of the mesophyll cells, accelerates the interconversion of CO2 to HCO3-, the substrate used by phosphoenolpyruvate carboxylase (PEPC) in the first step of C4 photosynthesis. Here we describe the identification and characterization of low CO2-responsive mutant 1 (lcr1) isolated from an N-nitroso-N-methylurea- (NMU) treated Setaria viridis mutant population. Forward genetic investigation revealed that the mutated gene Sevir.5G247800 of lcr1 possessed a single nucleotide transition from cytosine to thymine in a ß-CA gene causing an amino acid change from leucine to phenylalanine. This resulted in severe reduction in growth and photosynthesis in the mutant. Both the CO2 compensation point and carbon isotope discrimination values of the mutant were significantly increased. Growth of the mutants was stunted when grown under ambient pCO2 but recovered at elevated pCO2. Further bioinformatics analyses revealed that the mutation has led to functional changes in one of the conserved residues of the protein, situated near the catalytic site. CA transcript accumulation in the mutant was 80% lower, CA protein accumulation 30% lower, and CA activity ~98% lower compared with the wild type. Changes in the abundance of other primary C4 pathway enzymes were observed; accumulation of PEPC protein was significantly increased and accumulation of malate dehydrogenase and malic enzyme decreased. The reduction of CA protein activity and abundance in lcr1 restricts the supply of bicarbonate to PEPC, limiting C4 photosynthesis and growth. This study establishes Sevir.5G247800 as the major CA allele in Setaria for C4 photosynthesis and provides important insights into the function of CA in C4 photosynthesis that would be required to generate a rice plant with a functional C4 biochemical pathway.
Subject(s)
Carbonic Anhydrases , Photosynthesis , Plant Proteins , Setaria Plant , Carbon Dioxide , Carbonic Anhydrases/genetics , Carbonic Anhydrases/metabolism , Mesophyll Cells/metabolism , Setaria Plant/enzymology , Setaria Plant/geneticsABSTRACT
BACKGROUND: The purpose of the present study was to determine the airway changes in skeletal class II division 1 malocclusion patients with mandibular retrognathism, treated with Twin-Block (TB) appliance. METHODS: Airway assessment was carried for twelve patients (mean age 11.7 ± 1.1 years) who underwent myofunctional therapy using TB appliance for correction of skeletal class II division 1 malocclusion with mandibular retrognathism. Acoustic pharyngometry (AP) was used to assess and quantify the comparative changes in the upper airway, pretreatment and posttreatment. RESULTS: Data acquired was subjected to appropriate statistical analysis. The paired 't' test was used to compare pre-treatment (T0) and after the positive pterygoid response (T1). TB appliance increased mean minimum airway area by 0.28 ± 0.25 cm2 and mean airway by 0.47 ± 0.44 cm2 with 95% CI. Posttreatment minimum airway and mean area changes were found to be statistically significant (P-value<0.01). CONCLUSION: TB appliance therapy has a positive effect on upper airway and is beneficial for the treatment of sleep-related disorders associated with Class II division 1 malocclusion for achieving positive functional changes, esthetics, and healthier quality of life.
ABSTRACT
BACKGROUND: Promoter motifs in Entamoeba histolytica were earlier analysed using microarray data with lower dynamic range of gene expression. Additionally, previous transcriptomic studies did not provide information on the nature of highly transcribed genes, and downstream promoter motifs important for gene expression. To address these issues we generated RNA-Seq data and identified the high and low expressing genes, especially with respect to virulence potential. We analysed sequences both upstream and downstream of start site for important motifs. RESULTS: We used RNA-Seq data to classify genes according to expression levels, which ranged six orders of magnitude. Data were validated by reporter gene expression. Virulence-related genes (except AIG1) were amongst the highly expressed, while some kinases and BspA family genes were poorly expressed. We looked for conserved motifs in sequences upstream and downstream of the initiation codon. Following enrichment by AME we found seven motifs significantly enriched in high expression- and three in low expression-classes. Two of these motifs (M4 and M6) were located downstream of AUG, were exclusively enriched in high expression class, and were mostly found in ribosomal protein, and translation-related genes. Motif deletion resulted in drastic down regulation of reporter gene expression, showing functional relevance. Distribution of core promoter motifs (TATA, GAAC, and Inr) in all genes revealed that genes with downstream motifs were not preferentially associated with TATA-less promoters. We looked at gene expression changes in cells subjected to growth stress by serum starvation, and experimentally validated the data. Genes showing maximum up regulation belonged to the low or medium expression class, and included genes in signalling pathways, lipid metabolism, DNA repair, Myb transcription factors, BspA, and heat shock. Genes showing maximum down regulation belonged to the high or medium expression class. They included genes for signalling factors, actin, Ariel family, and ribosome biogenesis factors. CONCLUSION: Our analysis has added important new information about the E. histolytica transcriptome. We report for the first time two downstream motifs required for gene expression, which could be used for over expression of E. histolytica genes. Most of the virulence-related genes in this parasite are highly expressed in culture.
Subject(s)
Entamoeba histolytica/pathogenicity , Gene Expression Profiling/methods , Virulence Factors/genetics , Entamoeba histolytica/genetics , Gene Expression Regulation, Neoplastic , Gene Regulatory Networks , Promoter Regions, Genetic , Sequence Analysis, RNA , Whole Genome SequencingABSTRACT
Leaves are derived from heterotrophic meristem tissue that, at some point, must make the transition to autotrophy via the initiation of photosynthesis. However, the timing and spatial coordination of the molecular and cellular processes underpinning this switch are poorly characterized. Here, we report on the identification of a specific stage in rice (Oryza sativa) leaf development (P3/P4 transition) when photosynthetic competence is first established. Using a combined physiological and molecular approach, we show that elements of stomatal and vascular differentiation are coordinated with the onset of measurable light absorption for photosynthesis. Moreover, by exploring the response of the system to environmental perturbation, we show that the earliest stages of rice leaf development have significant plasticity with respect to elements of cellular differentiation of relevance for mature leaf photosynthetic performance. Finally, by performing an RNA sequencing analysis targeted at the early stages of rice leaf development, we uncover a palette of genes whose expression likely underpins the acquisition of photosynthetic capability. Our results identify the P3/P4 transition as a highly dynamic stage in rice leaf development when several processes for the initiation of photosynthetic competence are coordinated. As well as identifying gene targets for future manipulation of rice leaf structure/function, our data highlight a developmental window during which such manipulations are likely to be most effective.
Subject(s)
Chlorophyll/metabolism , Gene Expression Profiling/methods , Oryza/genetics , Photosynthesis/genetics , Plant Leaves/genetics , Chlorophyll/chemistry , Fluorescence , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , Light , Microscopy, Electron, Scanning , Microscopy, Electron, Transmission , Oryza/growth & development , Oryza/metabolism , Photosynthesis/radiation effects , Plant Leaves/growth & development , Plant Leaves/metabolism , Plant Stomata/genetics , Plant Stomata/metabolism , Plant Stomata/ultrastructure , Plant Vascular Bundle/genetics , Plant Vascular Bundle/growth & development , Plant Vascular Bundle/metabolism , Plastids/genetics , Plastids/metabolism , Plastids/ultrastructure , Time FactorsSubject(s)
COVID-19 , Female , Humans , Parturition , Pregnancy , Prospective Studies , Referral and Consultation , SARS-CoV-2ABSTRACT
The specification of vascular patterning in plants has interested plant biologists for many years. In the last decade a new context has emerged for this interest. Specifically, recent proposals to engineer C(4) traits into C(3) plants such as rice require an understanding of how the distinctive venation pattern in the leaves of C(4) plants is determined. High vein density with Kranz anatomy, whereby photosynthetic cells are arranged in encircling layers around vascular bundles, is one of the major traits that differentiate C(4) species from C(3) species. To identify genetic factors that specify C(4) leaf anatomy, we generated ethyl methanesulfonate- and γ-ray-mutagenized populations of the C(4) species sorghum (Sorghum bicolor), and screened for lines with reduced vein density. Two mutations were identified that conferred low vein density. Both mutations segregated in backcrossed F(2) populations as homozygous recessive alleles. Bulk segregant analysis using next-generation sequencing revealed that, in both cases, the mutant phenotype was associated with mutations in the CYP90D2 gene, which encodes an enzyme in the brassinosteroid biosynthesis pathway. Lack of complementation in allelism tests confirmed this result. These data indicate that the brassinosteroid pathway promotes high vein density in the sorghum leaf, and suggest that differences between C(4) and C(3) leaf anatomy may arise in part through differential activity of this pathway in the two leaf types.
Subject(s)
Brassinosteroids/metabolism , Cytochrome P-450 Enzyme System/genetics , Plant Leaves/genetics , Plant Proteins/genetics , Sorghum/genetics , Cytochrome P-450 Enzyme System/metabolism , Plant Leaves/metabolism , Plant Proteins/metabolism , Sorghum/metabolismABSTRACT
Comparison of transcriptome for candidate gene discovery has become an important tool for biologists. While such studies lack the degree of resolution one gets from well-designed forward or reverse genetic studies, nevertheless, this has been a method of choice for giving coarse insight into the underlying biological processes or mechanisms. This was further accelerated with the availability of sequencing technologies. While many pipelines are available for RNA-seq data analysis, the protocol discussed here will guide the first-time users for conducting routine RNA-seq analysis using whole genome sequence as reference.
Subject(s)
Gene Expression Profiling , RNA-Seq , Software , RNA-Seq/methods , Gene Expression Profiling/methods , Computational Biology/methods , Humans , Sequence Analysis, RNA/methods , Transcriptome , High-Throughput Nucleotide Sequencing/methodsABSTRACT
Apexification is a technique used to create a mineralized barrier in a root having an open apex or to continue the apical growth of a root that is still not fully developed in teeth with necrotic pulps. Biodentine is a versatile material which can help in achieving apical closure as well as reinforcement of thin dentinal walls, thus improving the overall prognosis and preserving the natural dentition. In the present case, the radiographic evaluation showed thinner dentinal walls and an open apex with periapical radiolucency in relation to the upper right central incisor. The large open root apex and thin dentin walls of immature permanent teeth render them challenging to treat with root canal therapy. This case report describes a modified single-step apexification procedure and strengthening of the dentinal walls of the root canal using Biodentine.
ABSTRACT
The draft genome sequence of an agriculturally important actinobacterial species Amycolatopsis sp. BCA-696 was developed and characterized in this study. Amycolatopsis BCA-696 is known for its biocontrol properties against charcoal rot and also for plant growth-promotion (PGP) in several crop species. The next-generation sequencing (NGS)-based draft genome of Amycolatopsis sp. BCA-696 comprised of ~ 9.05 Mb linear chromosome with 68.75% GC content. In total, 8716 protein-coding sequences and 61 RNA-coding sequences were predicted in the genome. This newly developed genome sequence has been also characterized for biosynthetic gene clusters (BGCs) and biosynthetic pathways. Furthermore, we have also reported that the Amycolatopsis sp. BCA-696 produces the glycopeptide antibiotic vancomycin that inhibits the growth of pathogenic gram-positive bacteria. A comparative analysis of the BCA-696 genome with publicly available closely related genomes of 14 strains of Amycolatopsis has also been conducted. The comparative analysis has identified a total of 4733 core and 466 unique orthologous genes present in the BCA-696 genome The unique genes present in BCA-696 was enriched with antibiotic biosynthesis and resistance functions. Genome assembly of the BCA-696 has also provided genes involved in key pathways related to PGP and biocontrol traits such as siderophores, chitinase, and cellulase production.
Subject(s)
Amycolatopsis , Genome, Bacterial , Genomics , Genomics/methods , Amycolatopsis/genetics , Amycolatopsis/metabolism , Multigene Family , Plant Development/genetics , High-Throughput Nucleotide Sequencing , Phylogeny , Vancomycin/pharmacologyABSTRACT
Sequencing technologies have rapidly evolved over the past two decades, and new technologies are being continually developed and commercialized. The emerging sequencing technologies target generating more data with fewer inputs and at lower costs. This has also translated to an increase in the number and type of corresponding applications in genomics besides enhanced computational capacities (both hardware and software). Alongside the evolving DNA sequencing landscape, bioinformatics research teams have also evolved to accommodate the increasingly demanding techniques used to combine and interpret data, leading to many researchers moving from the lab to the computer. The rich history of DNA sequencing has paved the way for new insights and the development of new analysis methods. Understanding and learning from past technologies can help with the progress of future applications. This review focuses on the evolution of sequencing technologies, their significant enabling role in generating plant genome assemblies and downstream applications, and the parallel development of bioinformatics tools and skills, filling the gap in data analysis techniques.
ABSTRACT
PREMISE OF THE STUDY: Next-generation sequencing (NGS) technologies are frequently used for resequencing and mining of single nucleotide polymorphisms (SNPs) by comparison to a reference genome. In crop species such as chickpea (Cicer arietinum) that lack a reference genome sequence, NGS-based SNP discovery is a challenge. Therefore, unlike probability-based statistical approaches for consensus calling and by comparison with a reference sequence, a coverage-based consensus calling (CbCC) approach was applied and two genotypes were compared for SNP identification. METHODS: A CbCC approach is used in this study with four commonly used short read alignment tools (Maq, Bowtie, Novoalign, and SOAP2) and 15.7 and 22.1 million Illumina reads for chickpea genotypes ICC4958 and ICC1882, together with the chickpea trancriptome assembly (CaTA). KEY RESULTS: A nonredundant set of 4543 SNPs was identified between two chickpea genotypes. Experimental validation of 224 randomly selected SNPs showed superiority of Maq among individual tools, as 50.0% of SNPs predicted by Maq were true SNPs. For combinations of two tools, greatest accuracy (55.7%) was reported for Maq and Bowtie, with a combination of Bowtie, Maq, and Novoalign identifying 61.5% true SNPs. SNP prediction accuracy generally increased with increasing reads depth. CONCLUSIONS: This study provides a benchmark comparison of tools as well as read depths for four commonly used tools for NGS SNP discovery in a crop species without a reference genome sequence. In addition, a large number of SNPs have been identified in chickpea that would be useful for molecular breeding.
Subject(s)
Cicer/genetics , Consensus Sequence , Crops, Agricultural/genetics , Genome, Plant , Polymorphism, Single Nucleotide , Sequence Analysis, DNA/methods , Base Sequence , Chromosome Mapping/methods , Computational Biology/methods , DNA, Plant/genetics , Genetic Variation , Genotype , Reference Standards , Reproducibility of Results , Sequence Alignment/methods , TranscriptomeABSTRACT
BACKGROUND: Several tools are available to identify miRNAs from deep-sequencing data, however, only a few of them, like miRDeep, can identify novel miRNAs and are also available as a standalone application. Given the difference between plant and animal miRNAs, particularly in terms of distribution of hairpin length and the nature of complementarity with its duplex partner (or miRNA star), the underlying (statistical) features of miRDeep and other tools, using similar features, are likely to get affected. RESULTS: The potential effects on features, such as minimum free energy, stability of secondary structures, excision length, etc., were examined, and the parameters of those displaying sizable changes were estimated for plant specific miRNAs. We found most of these features acquired a new set of values or distributions for plant specific miRNAs. While the length of conserved positions (nucleus) in mature miRNAs were relatively longer in plants, the difference in distribution of minimum free energy, between real and background hairpins, was marginal. However, the choice of source (species) of background sequences was found to affect both the minimum free energy and miRNA hairpin stability. The new parameters were tested on an Illumina dataset from maize seedlings, and the results were compared with those obtained using default parameters. The newly parameterized model was found to have much improved specificity and sensitivity over its default counterpart. CONCLUSIONS: In summary, the present study reports behavior of few general and tool-specific statistical features for improving the prediction accuracy of plant miRNAs from deep-sequencing data.
Subject(s)
Computational Biology/methods , MicroRNAs/chemistry , Plants/genetics , RNA, Plant/chemistry , Data Interpretation, Statistical , Nucleic Acid ConformationABSTRACT
Single feature polymorphisms (SFPs) are microarray-based molecular markers that are detected by hybridization of DNA or cRNA to oligonucleotide probes. With an objective to identify the potential polymorphic markers for drought tolerance in pigeonpea [Cajanus cajan (L.) Millspaugh], an important legume crop for the semi-arid tropics but deficient in genomic resources, Affymetrix Genome Arrays of soybean (Glycine max), a closely related species of pigeonpea were used on cRNA of six parental genotypes of three mapping populations of pigeonpea segregating for agronomic traits like drought tolerance and pod borer (Helicoverpa armigiera) resistance. By using robustified projection pursuit method on 15 pair-wise comparisons for the six parental genotypes, 5,692 SFPs were identified. Number of SFPs varied from 780 (ICPL 8755 × ICPL 227) to 854 (ICPL 151 × ICPL 87) per parental combination of the mapping populations. Randomly selected 179 SFPs were used for validation by Sanger sequencing and good quality sequence data were obtained for 99 genes of which 75 genes showed sequence polymorphisms. While associating the sequence polymorphisms with SFPs detected, true positives were observed for 52.6% SFPs detected. In terms of parental combinations of the mapping populations, occurrence of true positives was 34.48% for ICPL 151 × ICPL 87, 41.86% for ICPL 8755 × ICPL 227, and 81.58% for ICP 28 × ICPW 94. In addition, a set of 139 candidate genes that may be associated with drought tolerance has been identified based on gene ontology analysis of the homologous pigeonpea genes to the soybean genes that detected SFPs between the parents of the mapping populations segregating for drought tolerance.
Subject(s)
Cajanus/genetics , Dehydration/genetics , Polymorphism, Genetic , Cajanus/physiology , Droughts , Gene Expression Profiling , Genetic Markers , Genome, Plant , Genotype , Oligonucleotide Array Sequence Analysis , Stress, Physiological/geneticsABSTRACT
Development of insect resistance to biopesticides is a current and pertinent global issue. Earlier, it was established that lepidopteran larvae can recover from Bt intoxication via a midgut regenerative response and subsequently generate resistance. Molecular aspects of restoration of the midgut integrity following toxin exposure are emerging recently. In the present study, we bring out the pivotal role of gut arylphorin in mediating the midgut regenerative response following sublethal Bt exposure in Achaea janata. Bt-induced midgut damage was characterized by microscopic analysis using differential interference contrast (DIC) and immunofluorescence (IF). Extensive disruption of brush-border membrane, associated with the underlying cytoskeletal alterations including F-actin, α-actin and ß-tubulin was observed. Single-photon fluorescence microscopy combined with fluorescence lifetime imaging (FLIM) established the metabolic state associated with enhanced stem cell proliferation and migration from the basal side towards the luminal side following the damage. In-silico analysis revealed the phylogenetic relationship of gut arylphorin with closely related insect species and indicated the presence of two different subunits. Transient RNAi knockdown of the arylphorin resulted in diminished expression of mitotic Cyclin B mRNA levels. Human monoclonal Cyclin B antibody cross-reactivity with the Cyclin B located in the stem cells further validate the role of arylphorin as the mitogenic factor responsible for stem cell proliferation and epithelial regeneration. An in-depth understanding of resistance mechanisms will aid in the design of new strategies for the long-term usage and efficacy of Bt technology against pest control.