Genome-wide identification of the MIOX gene family and their expression profile in cotton development and response to abiotic stress

The enzyme myo-inositol oxygenase (MIOX) catalyzes the myo-inositol into glucuronic acid. In this study, 6 MIOX genes were identified from all of the three diploid cotton species (Gossypium arboretum, Gossypium herbaceum and Gossypium raimondii) and Gossypioides kirkii, 12 MIOX genes were identified from two domesticated tetraploid cottons Gossypium hirsutum, Gossypium barbadense, and 11 MIOX genes were identified from three wild tetraploid cottons Gossypium tomentosum, Gossypium mustelinum and Gossypium darwinii. The number of MIOX genes in tetraploid cotton genome is roughly twice that of diploid cotton genome. Members of MIOX family were classified into six groups based on the phylogenetic analysis. Integrated analysis of collinearity events and chromosome locations suggested that both whole genome duplication and segmental duplication events contributed to the expansion of MIOX genes during cotton evolution. The ratios of non-synonymous (Ka) and synonymous (Ks) substitution rates revealed that purifying selection was the main force driving the evolution of MIOX genes. Numerous cis-acting elements related to light responsive element, defense and stress responsive element were identified in the promoter of the MIOX genes. Expression analyses of MIOX genes based on RNA-seq data and quantitative real time PCR showed that MIOX genes within the same group shared similar expression patterns with each other. All of these results provide the foundation for further study of the biological functions of MIOX genes in cotton environmental adaptability.

Plant regeneration via somatic embryogenesis in diploid cultivated cotton (Gossypium arboreum L.)

The cultivated diploid cotton species G. arboreum offers a better opportunity to elucidate gene structure and function compared to the allotetraploid cotton species through genetic transformation, the reliable and efficient method for high frequency somatic embryogenesis and plant regeneration in G. arboreum is urgent need to be established. Callus was induced from hypocotyl, root and cotyledon of G. arboreum seedlings on MSB (MS salts and B5 vitamins) medium with 0.09 µM 2,4-D and 2.32 µM KT. The embryogenic callus was induced on MS5 medium from the suspended cultures of several cycles of alternate liquid-solid culture, which was critical step for somatic embryogenesis. The liquid medium of MS4 was supplemented with 0.1g/L NaCl, 0.1g/L KCl and 0.1g/L CuSO4. The solid medium of MS5 for embryogenic callus effective induction was supplemented with 37.59 mM KNO3 + 62.47 µM NH4NO3 and 2.46 µM IBA + 0.93 µM KT or 0.045 µM 2,4-D + 2.46 µM IBA + 0.465 µM KT. During callus growing on different media, callus was effectively selected for subculture or treatment according to cell morphology to induce embryogenic callus and somatic embryos. Somatic embryo maturation and germination were better on MS5 medium with maltose or glucose + maltose than the single glucose. The regenerated plantlets with well-developed roots were directly transferred to soil or grafted onto the germinated cotton plantlets. The feasible process of plant regeneration via somatic embryogenesis in diploid cultivated species was established and needed to be improved and optimized for the gene functional analysis and gene editing in the diploid cotton species.

Genome-Wide Identification and Characterization of The MIOX Gene Family in Cotton

Background: The enzyme myo-inositol oxygenase (MIOX) catalyzes the myo-inositol into glucuronic acid. Previous studies indicated that MIOX may play an important role in plant responses to abiotic stresses. Cotton is a major source of natural and renewable textile. However, the MIOX gene family of cotton has not been systematically identified and characterized yet.Results: In this study, 6 MIOX genes were identified from all of the three diploid cotton species (Gossypium arboretum, Gossypium herbaceum and Gossypium raimondii), 12 MIOX genes were identified from two domesticated tetraploid cottons Gossypium hirsutum, Gossypium barbadense, and 11 MIOX genes were identified from three wild tetraploid cottons Gossypium tomentosum, Gossypium mustelinum and Gossypium darwinii. The number of MIOX genes in tetraploid cotton genome is roughly twice that of diploid cotton genome. Members of MIOX family were classified into six groups based on the phylogenetic analysis. Integrated analysis of collinearity events and chromosome locations suggested that both whole genome duplication and segmental duplication events contributed to the expansion of MIOX genes during cotton evolution. The ratios of non-synonymous (Ka) and synonymous (Ks) substitution rates revealed that purifying selection was the main force driving the evolution of MIOX genes. Numerous cis-acting elements related to light responsive element, defense and stress responsive element were identified in the promoter of the MIOX genes. Expression analyses of MIOX genes based on RNA-seq data showed that MIOX genes within the same group shared similar expression patterns with each other.Conclusions: In this work, we systematically analyzed MIOX genes from eight Gossypium genomes and the Gossypioides kirkii genome using a set of bioinformatics approaches. All of these results provide the foundation for further study of the biological functions of MIOX genes in cotton environmental adaptability.

Morphological description of a novel synthetic allotetraploid(A1A1G3G3) of Gossypium herbaceum L.and G.nelsonii Fryx. suitable for disease-resistant breeding applications

Wild species of Gossypium ssp. are an important source of traits for improving commercial cotton cultivars. Previous reports show that Gossypium herbaceum L. and Gossypium nelsonii Fryx. have better disease resistance characteristics than commercial cotton varieties. However, chromosome ploidy and biological isolation make it difficult to hybridize diploid species with the tetraploid Gossypium hirsutum L. We developed a new allotetraploid cotton genotype (A1A1G3G3) using a process of distant hybridization within wild cotton species to create new germplasms. First of all, G. herbaceum and G. nelsonii were used for interspecific hybridization to obtain F1 generation. Afterwards, apical meristems of the F1 diploid cotton plants were treated with colchicine to induce chromosome doubling. The new interspecific F1 hybrid and S1 cotton plants originated from chromosome duplication, were tested via morphological and molecular markers and confirmed their tetraploidy through flowrometric and cytological identification. The S1 tetraploid cotton plants was crossed with a TM-1 line and fertile hybrid offspring were obtained. These S2 offsprings were tested for resistance to Verticillium wilt and demonstrated adequate tolerance to this fungi. The results shows that the new S1 cotton line could be used as parental material for hybridization with G. hirsutum to produce pathogen-resistant cotton hybrids. This new S1 allotetraploid genotype will contributes to the enrichment of Gossypium germplasm resources and is expected to be valuable in polyploidy evolutionary studies.

Assessment of genetic divergence in diploid cotton (Gossypium arboreum L.) germplasm using fibre quality traits

Cotton is one of the most important crops among natural fibres. Fibre quality determines the spinning ability, which is negatively correlated with yield and yield-contributing traits. Limited efforts have been made to improve fibre quality and yield in diploid cotton. Therefore, screening a large panel of germplasm lines can help identify genotypes with better fibre quality and yield. We evaluated 712 desi cotton genotypes for fibre quality traits. The genotypes showed a significant difference for all the traits, suggesting considerable variability for fibre quality improvement. Fibre length and strength showed high phenotypic and genotypic coefficients of variation. Heritability was high for fibre strength, length, and elongation. Fibre length and strength were positively correlated; however, micronaire was negatively correlated with these two traits. Superior accessions were identified for fibre length (11), strength (20), uniformity (7), and elongation (25) among genotypes. Most of the desi lines (71%) had medium micronaire values. Twenty accessions identified were ideal for spinning, showing the fibre strength-to-length ratio of one. Cluster analysis based on Euclidean distance grouped all 712 accessions into four major clusters. Principal component analysis biplot revealed that accessions AC3418, 360-SP1, AC3522B, Kanpur A, Gao16CB-9, and AC3370 were genetically diverse. The superior accessions for fibre quality identified in this study are potential lines for the diploid cotton improvement programme.

Full-length annotation with multi-strategy RNA-seq uncovers transcriptional regulation of lncRNAs in diploid cotton G. arboreum1

Long noncoding RNAs (lncRNAs) are crucial factors during plant development and environmental responses. High-throughput and accurate identification of lncRNAs is still lacking in plants. To build an accurate atlas of lncRNA in cotton, we combined Isoform-sequencing (Iso-seq), strand-specific RNA-seq (ssRNA-seq), cap analysis gene expression (CAGE-seq) with PolyA-seq and compiled a pipeline named plant full-length lncRNA (PULL) to integrate multi-omics data. A total of 9240 lncRNAs from 21 tissue samples of the diploid cotton Gossypium arboreum were identified. We revealed that alternative usage of transcription start site (TSS) and transcription end site (TES) of lncRNAs occurs pervasively during plant growth and responses to stress. We identified the lncRNAs which co-expressed or be linked to the protein coding genes (PCGs) or GWAS studied SNPs associated with ovule and fiber development. We also mapped the genome-wide binding sites of two lncRNAs with chromatin isolation by RNA purification sequencing (ChIRP-seq) and validated the trans transcriptional regulation of lnc-Ga13g0352 via virus induced gene suppression (VIGS) assay. These findings provide valuable research resources for plant community and broaden our understandings of biogenesis and regulation function of plant lncRNAs.One sentence summaryThe full-length annotation and transcriptional regulation of long noncoding RNAs in cotton.