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Isolation and expression analysis of EcbZIP17 from different finger millet genotypes shows conserved nature of the gene.

ABSTRACT: Basic leucine zipper (bZIP) transcription factors comprise one of the largest gene families in plants. They play a key role in almost every aspect of plant growth and development and also in biotic and abiotic stress tolerance. In this study, we report isolation and characterization of EcbZIP17, a group B bZIP transcription factor from a climate smart cereal, finger millet (Eleusine coracana L.). The genomic sequence of EcbZIP17 is 2662 bp long encompassing two exons and one intron with ORF of 1722 bp and peptide length of 573 aa. This gene is homologous to AtbZIP17 (Arabidopsis), ZmbZIP17 (maize) and OsbZIP60 (rice) which play a key role in endoplasmic reticulum (ER) stress pathway. In silico analysis confirmed the presence of basic leucine zipper (bZIP) and transmembrane (TM) domains in the EcbZIP17 protein. Allele mining of this gene in 16 different genotypes by Sanger sequencing revealed no variation in nucleotide sequence, including the 618 bp long intron. Expression analysis of EcbZIP17 under heat stress exhibited similar pattern of expression in all the genotypes across time intervals with highest upregulation after 4 h. The present study established the conserved nature of EcbZIP17 at nucleotide and expression level.

SUBMITTER: Chopperla R

PROVIDER: S-EPMC5608649 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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Isolation and expression analysis of EcbZIP17 from different finger millet genotypes shows conserved nature of the gene.

Chopperla Ramakrishna R Singh Sonam S Mohanty Sasmita S Reddy Nanja N Padaria Jasdeep C JC Solanke Amolkumar U AU

3 Biotech 20170921 5

Basic leucine zipper (bZIP) transcription factors comprise one of the largest gene families in plants. They play a key role in almost every aspect of plant growth and development and also in biotic and abiotic stress tolerance. In this study, we report isolation and characterization of EcbZIP17, a group B bZIP transcription factor from a climate smart cereal, finger millet (Eleusine coracana L.). The genomic sequence of EcbZIP17 is 2662 bp long encompassing two exons and one ...[more]

PMID: 28955639

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Project description:Eleusine coracana, finger millet, is a multipurpose crop cultivated in arid and semi-arid regions of Africa and Asia. RNA sequencing (RNA-seq) was used in this study to obtain valuable genomic resources and identify genes differentially expressed between Al-tolerant and Al-susceptible genotypes. Two groups of finger millet genotypes were used: Al-tolerant (215836, 215845, and 229722) and Al-susceptible (212462, 215804 and 238323). The analysis of the RNA-seq data resulted in 198,546 unigenes, 56.5% of which were annotated with significant hits in one or more of the following six databases: NR (48.8%), GO (29.7%), KEGG (45%), PlantTFDB (19.0%), Uniprot (49.2%), and NT (46.2%). It is noteworthy that only 220 unigenes in the NR database had significant hits against finger millet sequences suggesting that finger millet's genomic resources are scarce. The gene expression analysis revealed that 322 genes were significantly differentially expressed between the Al-tolerant and Al-susceptible genotypes, of which 40.7% were upregulated while 59.3% were downregulated in Al-tolerant genotypes. Among the significant DEGs, 54.7% were annotated in the GO database with the top hits being ATP binding (GO:0005524) and DNA binding (GO:0003677) in the molecular function, DNA integration (GO:0015074) and cell redox homeostasis in the biological process, as well as cellular anatomical entity and intracellular component in the cellular component GO classes. Several of the annotated DEGs were significantly enriched for their corresponding GO terms. The KEGG pathway analysis resulted in 60 DEGs that were annotated with different pathway classes, of which carbohydrate metabolism and signal transduction were the most prominent. The homologs of a number of significant DEGs have been previously reported as being associated with Al or other abiotic stress responses in various crops, including carboxypeptidase SOL1, HMA3, AP2, bZIP, C3H, and WRKY TF genes. A more detailed investigation of these and other DEGs will enable genomic-led breeding for Al tolerance in finger millet. RNA-seq data analysis also yielded 119,073 SNP markers, the majority of which had PIC values above 0.3, indicating that they are highly informative. Additionally, 3,553 single-copy SSR markers were identified, of which trinucleotide SSRs were the most prevalent. These genomic resources contribute substantially to the enrichment of genomic databases for finger millet, and facilitate future research on this crop.

Project description:BackgroundIn finger millet, calcium is one of the important and abundant mineral elements. The molecular mechanisms involved in calcium accumulation in plants remains poorly understood. Transcriptome sequencing of genetically diverse genotypes of finger millet differing in grain calcium content will help in understanding the trait.Principal findingIn this study, the transcriptome sequencing of spike tissues of two genotypes of finger millet differing in their grain calcium content, were performed for the first time. Out of 109,218 contigs, 78 contigs in case of GP-1 (Low Ca genotype) and out of 120,130 contigs 76 contigs in case of GP-45 (High Ca genotype), were identified as calcium sensor genes. Through in silico analysis all 82 unique calcium sensor genes were classified into eight calcium sensor gene family viz., CaM & CaMLs, CBLs, CIPKs, CRKs, PEPRKs, CDPKs, CaMKs and CCaMK. Out of 82 genes, 12 were found diverse from the rice orthologs. The differential expression analysis on the basis of FPKM value resulted in 24 genes highly expressed in GP-45 and 11 genes highly expressed in GP-1. Ten of the 35 differentially expressed genes could be assigned to three documented pathways involved mainly in stress responses. Furthermore, validation of selected calcium sensor responder genes was also performed by qPCR, in developing spikes of both genotypes grown on different concentration of exogenous calcium.ConclusionThrough de novo transcriptome data assembly and analysis, we reported the comprehensive identification and functional characterization of calcium sensor gene family. The calcium sensor gene family identified and characterized in this study will facilitate in understanding the molecular basis of calcium accumulation and development of calcium biofortified crops. Moreover, this study also supported that identification and characterization of gene family through Illumina paired-end sequencing is a potential tool for generating the genomic information of gene family in non-model species.

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Isolation and expression analysis of EcbZIP17 from different finger millet genotypes shows conserved nature of the gene.

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Isolation and expression analysis of <i>EcbZIP17</i> from different finger millet genotypes shows conserved nature of the gene.

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