Evolutionary divergence of duplicated genomes in newly described allotetraploid cottons.

Allotetraploid cotton (Gossypium) species represents a model system for the study of plant polyploidy, molecular evolution, and domestication. Here, chromosome-scale genome sequences were obtained and assembled for two recently described wild species of tetraploid cotton, Gossypium ekmanianum [(AD)6, Ge] and Gossypium stephensii [(AD)7, Gs], and one early form of domesticated Gossypium hirsutum, race punctatum [(AD)1, Ghp]. Based on phylogenomic analysis, we provide a dated whole-genome level perspective for the evolution of the tetraploid Gossypium clade and resolved the evolutionary relationships of Gs, Ge, and domesticated G. hirsutum. We describe genomic structural variation that arose during Gossypium evolution and describe its correlates-including phenotypic differentiation, genetic isolation, and genetic convergence-that contributed to cotton biodiversity and cotton domestication. Presence/absence variation is prominent in causing cotton genomic structural variations. A presence/absence variation-derived gene encoding a phosphopeptide-binding protein is implicated in increasing fiber length during cotton domestication. The relatively unimproved Ghp offers the potential for gene discovery related to adaptation to environmental challenges. Expanded gene families enoyl-CoA δ isomerase 3 and RAP2-7 may have contributed to abiotic stress tolerance, possibly by targeting plant hormone-associated biochemical pathways. Our results generate a genomic context for a better understanding of cotton evolution and for agriculture.

Genome-wide analyses of member identification, expression pattern, and protein-protein interaction of EPF/EPFL gene family in Gossypium.

BACKGROUND: Epidermal patterning factor / -like (EPF/EPFL) gene family encodes a class of cysteine-rich secretory peptides, which are widelyfound in terrestrial plants.Multiple studies has indicated that EPF/EPFLs might play significant roles in coordinating plant development and growth, especially as the morphogenesis processes of stoma, awn, stamen, and fruit skin. However, few research on EPF/EPFL gene family was reported in Gossypium. RESULTS: We separately identified 20 G. raimondii, 24 G. arboreum, 44 G. hirsutum, and 44 G. barbadense EPF/EPFL genes in the 4 representative cotton species, which were divided into four clades together with 11 Arabidopsis thaliana, 13 Oryza sativa, and 17 Selaginella moellendorffii ones based on their evolutionary relationships. The similar gene structure and common motifs indicated the high conservation among the EPF/EPFL members, while the uneven distribution in chromosomes implied the variability during the long-term evolutionary process. Hundreds of collinearity relationships were identified from the pairwise comparisons of intraspecifc and interspecific genomes, which illustrated gene duplication might contribute to the expansion of cotton EPF/EPFL gene family. A total of 15 kinds of cis-regulatory elements were predicted in the promoter regions, and divided into three major categories relevant to the biological processes of development and growth, plant hormone response, and abiotic stress response. Having performing the expression pattern analyses with the basic of the published RNA-seq data, we found most of GhEPF/EPFL and GbEPF/EPFL genes presented the relatively low expression levels among the 9 tissues or organs, while showed more dramatically different responses to high/low temperature and salt or drought stresses. Combined with transcriptome data of developing ovules and fibers and quantitative Real-time PCR results (qRT-PCR) of 15 highly expressed GhEPF/EPFL genes, it could be deduced that the cotton EPF/EPFL genes were closely related with fiber development. Additionally, the networks of protein-protein interacting among EPF/EPFLs concentrated on the cores of GhEPF1 and GhEPF7, and thosefunctional enrichment analyses indicated that most of EPF/EPFLs participate in the GO (Gene Ontology) terms of stomatal development and plant epidermis development, and the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways of DNA or base excision repair. CONCLUSION: Totally, 132 EPF/EPFL genes were identified for the first time in cotton, whose bioinformatic analyses of cis-regulatory elements and expression patterns combined with qRT-PCR experiments to prove the potential functions in the biological processes of plant growth and responding to abiotic stresses, specifically in the fiber development. These results not only provide comprehensive and valuable information for cotton EPF/EPFL gene family, but also lay solid foundation for screening candidate EPF/EPFL genes in further cotton breeding.

Antibacterial activity and mechanisms of D-3263 against Staphylococcus aureus.

Multi-drug-resistant Staphylococcus aureus infections necessitate novel antibiotic development. D-3263, a transient receptor potential melastatin member 8 (TRPM8) agonist, has potential antineoplastic properties. Here, we reported the antibacterial and antibiofilm activities of D-3263. Minimum inhibitory concentrations (MICs) against S. aureus, Enterococcus faecalis and E. faecium were ≤ 50 µM. D-3263 exhibited bactericidal effects against clinical methicillin-resistant S. aureus (MRSA) and E. faecalis strains at 4× MIC. Subinhibitory D-3263 concentrations effectively inhibited S. aureus and E. faecalis biofilms, with higher concentrations also clearing mature biofilms. Proteomic analysis revealed differential expression of 29 proteins under 1/2 × MIC D-3263, influencing amino acid biosynthesis and carbohydrate metabolism. Additionally, D-3263 enhanced membrane permeability of S. aureus and E. faecalis. Bacterial membrane phospholipids phosphatidylethanolamine (PE), phosphatidylglycerol (PG), and cardiolipin (CL) dose-dependently increased D-3263 MICs. Overall, our data suggested that D-3263 exhibited potent antibacterial and antibiofilm activities against S. aureus by targeting the cell membrane.

Genome-wide chromatin accessibility landscape and dynamics of transcription factor networks during ovule and fiber development in cotton.

BACKGROUND: The development of cotton fiber is regulated by the orchestrated binding of regulatory proteins to cis-regulatory elements associated with developmental genes. The cis-trans regulatory dynamics occurred throughout the course of cotton fiber development are elusive. Here we generated genome-wide high-resolution DNase I hypersensitive sites (DHSs) maps to understand the regulatory mechanisms of cotton ovule and fiber development. RESULTS: We generated DNase I hypersensitive site (DHS) profiles from cotton ovules at 0 and 3 days post anthesis (DPA) and fibers at 8, 12, 15, and 18 DPA. We obtained a total of 1185 million reads and identified a total of 199,351 DHSs through ~ 30% unique mapping reads. It should be noted that more than half of DNase-seq reads mapped multiple genome locations and were not analyzed in order to achieve a high specificity of peak profile and to avoid bias from repetitive genomic regions. Distinct chromatin accessibilities were observed in the ovules (0 and 3 DPA) compared to the fiber elongation stages (8, 12, 15, and 18 DPA). Besides, the chromatin accessibility during ovules was particularly elevated in genomic regions enriched with transposable elements (TEs) and genes in TE-enriched regions were involved in ovule cell division. We analyzed cis-regulatory modules and revealed the influence of hormones on fiber development from the regulatory divergence of transcription factor (TF) motifs. Finally, we constructed a reliable regulatory network of TFs related to ovule and fiber development based on chromatin accessibility and gene co-expression network. From this network, we discovered a novel TF, WRKY46, which may shape fiber development by regulating the lignin content. CONCLUSIONS: Our results not only reveal the contribution of TEs in fiber development, but also predict and validate the TFs related to fiber development, which will benefit the research of cotton fiber molecular breeding.

Fine mapping and candidate gene analysis of qFL-A12-5: a fiber length-related QTL introgressed from Gossypium barbadense into Gossypium hirsutum.

KEY MESSAGE: The fiber length-related qFL-A12-5 identified in CSSLs introgressed from Gossypium barbadense into Gossypium hirsutum was fine-mapped to an 18.8 kb region on chromosome A12, leading to the identification of the GhTPR gene as a potential regulator of cotton fiber length. Fiber length is a key determinant of fiber quality in cotton, and it is a key target of artificial selection for breeding and domestication. Although many fiber length-related quantitative trait loci have been identified, there are few reports on their fine mapping or candidate gene validation, thus hampering efforts to understand the mechanistic basis of cotton fiber development. Our previous study identified the qFL-A12-5 associated with superior fiber quality on chromosome A12 in the chromosome segment substitution line (CSSL) MBI7747 (BC4F3:5). A single segment substitution line (CSSL-106) screened from BC6F2 was backcrossed to construct a larger segregation population with its recurrent parent CCRI45, thus enabling the fine mapping of 2852 BC7F2 individuals using denser simple sequence repeat markers to narrow the qFL-A12-5 to an 18.8 kb region of the genome, in which six annotated genes were identified in Gossypium hirsutum. Quantitative real-time PCR and comparative analyses led to the identification of GH_A12G2192 (GhTPR) encoding a tetratricopeptide repeat-like superfamily protein as a promising candidate gene for qFL-A12-5. A comparative analysis of the protein-coding regions of GhTPR among Hai1, MBI7747, and CCRI45 revealed two non-synonymous mutations. The overexpression of GhTPR resulted in longer roots in Arabidopsis, suggesting that GhTPR may regulate cotton fiber development. These results provide a foundation for future efforts to improve cotton fiber length.

Overexpression of cotton GhNAC072 gene enhances drought and salt stress tolerance in transgenic Arabidopsis.

BACKGROUND: Crops face several environmental stresses (biotic and abiotic), thus resulting in severe yield losses. Around the globe abiotic stresses are the main contributors of plant damages, primarily drought and salinity. Many genes and transcription factors are involved in abiotic and biotic stress responses. NAC TF (Transcription Factors) improves tolerance to stresses by controlling the physiological and enzyme activities of crops. RESULTS: In current research, GhNAC072 a highly upregulated TF in RNA-Seq was identified as a hub gene in the co-expression network analysis (WGCNA). This gene was transformed to Arabidopsis thaliana to confirm its potential role in drought and salt stress tolerance. Significant variations were observed in the morpho-physiological traits with high relative leaf water contents, chlorophyll contents, higher germination and longer root lengths of the overexpressed lines and low excised leaf loss and ion leakage as compared to the wildtype plants. Besides, overexpressed lines have higher amounts of antioxidants and low oxidant enzyme activities than the wildtype during the period of stress exposure. CONCLUSIONS: In summary, the above analysis showed that GhNAC072 might be the true candidate involved in boosting tolerance mechanisms under drought and salinity stress.

Homoeolog gene expression analysis reveals novel expression biases in upland hybrid cotton under intraspecific hybridization.

Hybridization is useful to enhance the yield potential of agronomic crops in the world. Cotton has genome doubling due to the allotetraploid process and hybridization in coordination with duplicated genome can produce more yield and adaptability. Therefore, the expression of homoeologous gene pairs between hybrids and inbred parents is vital to characterize the genetic source of heterosis in cotton. Investigation results of homoeolog gene pairs between two contrasting hybrids and their respective inbred parents identified 36853 homoeolog genes in hybrids. It was observed both high and low hybrids had similar trends in homoeolog gene expression patterns in each tissue under study. An average of 96% of homoeolog genes had no biased expression and their expressions were derived from the equal contribution of both parents. Besides, very few homoeolog genes (an average of 1%) showed no biased or novel expression in both hybrids. The functional analysis described secondary metabolic pathways had a majority of novel biased homoeolog genes in hybrids. These results contribute preliminary knowledge about how hybridization affects expression patterns of homoeolog gene pairs in upland cotton hybrids. Our study also highlights the functional genomics of metabolic genes to explore the genetic mechanism of heterosis in cotton.

Protoplast Dissociation and Transcriptome Analysis Provides Insights to Salt Stress Response in Cotton.

As one of the pioneer crops widely planted in saline-alkaline areas, Gossypium provides daily necessities, including natural fiber, vegetable proteins, and edible oils. However, cotton fiber yield and quality are highly influenced by salt stress. Therefore, elucidating the molecular mechanisms of cotton in response to salinity stress is importance to breed new cultivars with high tolerance. In this study, we first developed a method for single-cell RNA-seq based on isolating protoplast from cotton root tips; then, we studied the impact of salinity stress on gene expression profiling and their dynamic changes using the developed high-efficiency method for protoplast dissociation suitable for single-cell RNA-seq. A total of 3391 and 2826 differentially expressed genes (DEGs) were identified in salt-treated samples before and after protoplast dissociation, respectively, which were enriched into several molecular components, including response to stimulus, response to stress, and cellular macromolecule metabolic process by gene ontology (GO) analysis. Plant hormone signal transduction, phenylpropanoid biosynthesis, and MAPK signaling pathway were found to be enriched via Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Twenty-two and nine salinity-responsive DEGs participated in plant hormone signaling and MAPK signaling in roots, before and after protoplast dissociation, respectively; six upregulated DEGs were involved in ABA signaling transduction, namely, Ga04G2111, Ga07G0142, Ga09G2061, Ga10G0262, Ga01G0063, and Ga08G1915 which indicates their potential functions on plants adapting to salt stress. Additionally, 384 and 257 transcription factors (TFs) were differentially expressed in salt-stress roots before and after protoplast dissociation, respectively, of which significantly up-regulated TFs mainly belonged to the AP2/ERF-ERF family, which implied their potential roles responding to salt stress. These results not only provide novel insights to reveal the regulatory networks in plant's root response to salt stress, but also lay the solid foundation for further exploration on cellular heterogeneity by single-cell transcriptome sequencing.

Micro-FTIR combined with curve fitting method to study cellulose crystallinity of developing cotton fibers.

This study aimed to use micro-FTIR with transmission mode to investigate cellulose crystallinity of developing cotton fibers. Compared with ATR-FTIR method, we found that micro-FTIR can obtain more information of cellulose inside of the developing cotton fibers, especially in high wavenumber of 2800-3000 cm-1 region. Combined with curve fitting method, a new IR crystallinity index (CI) method named wax crystallinity index (WCI) was introduced to evaluate the cellulose crystallinity in the development of cotton fibers based on the peak and area ratios of 2900 cm-1/2850 cm-1 and 2900 cm-1/2920 cm-1. The obtained WCI values demonstrated an excellent coefficient of determination with X-ray diffraction (XRD) CI method with the value up to 0.99. This study suggested that micro-FTIR was an effective technique to qualitatively analyze the crystallinity in developing cotton fibers combined with curve fitting method.

High throughput deep sequencing elucidates the important role of lncRNAs in Foxtail millet response to herbicides.

Long non-coding RNAs (lncRNAs) play an important function in plant growth and development as well as response to stresses. However, little information was known in foxtail millet; no study was reported on lncRNAs in plant response to herbicide treatment. In this study, by using deep sequencing and advanced bioinformatic analysis, a total of 2547 lncRNAs were identified, including 787 known and 1760 novel lncRNAs. These lncRNAs are distributed across all 9 chromosomes, and the majority were located in the intergenic region with 1-2 exons. These lncRNAs were differentially expressed between different genotypes under different herbicide treatments. lncRNAs regulate plant growth and development as well as response to herbicide treatments through targeting protein-coding genes that directly relate to chemical metabolism and defense system. Multiple potential target genes and lncRNA-mRNA-miRNA gene networks were discovered. These results elucidate the potential roles of lncRNAs in plant response to herbicides.

Transcriptomic and proteomic analyses of a new cytoplasmic male sterile line with a wild Gossypium bickii genetic background.

BACKGROUND: Cotton is an important fiber crop but has serious heterosis effects, and cytoplasmic male sterility (CMS) is the major cause of heterosis in plants. However, to the best of our knowledge, no studies have investigated CMS Yamian A in cotton with the genetic background of Australian wild Gossypium bickii. Conjoint transcriptomic and proteomic analysis was first performed between Yamian A and its maintainer Yamian B. RESULTS: We detected 550 differentially expressed transcript-derived fragments (TDFs) and at least 1013 proteins in anthers at various developmental stages. Forty-two TDFs and 11 differentially expressed proteins (DEPs) were annotated by analysis in the genomic databases of G. austral, G. arboreum and G. hirsutum. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses were performed to better understand the functions of these TDFs and DEPs. Transcriptomic and proteomic results showed that UDP-glucuronosyl/UDP-glucosyltransferase, 60S ribosomal protein L13a-4-like, and glutathione S-transferase were upregulated; while heat shock protein Hsp20, ATPase, F0 complex, and subunit D were downregulated at the microspore abortion stage of Yamian A. In addition, several TDFs from the transcriptome and several DEPs from the proteome were detected and confirmed by quantitative real-time PCR as being expressed in the buds of seven different periods of development. We established the databases of differentially expressed genes and proteins between Yamian A and its maintainer Yamian B in the anthers at various developmental stages and constructed an interaction network based on the databases for a comprehensive understanding of the mechanism underlying CMS with a wild cotton genetic background. CONCLUSION: We first analyzed the molecular mechanism of CMS Yamian A from the perspective of omics, thereby providing an experimental basis and theoretical foundation for future research attempting to analyze the abortion mechanism of new CMS with a wild Gossypium bickii background and to realize three-line matching.

Genome sequencing of the Australian wild diploid species Gossypium australe highlights disease resistance and delayed gland morphogenesis.

The diploid wild cotton species Gossypium australe possesses excellent traits including resistance to disease and delayed gland morphogenesis, and has been successfully used for distant breeding programmes to incorporate disease resistance traits into domesticated cotton. Here, we sequenced the G. australe genome by integrating PacBio, Illumina short read, BioNano (DLS) and Hi-C technologies, and acquired a high-quality reference genome with a contig N50 of 1.83 Mb and a scaffold N50 of 143.60 Mb. We found that 73.5% of the G. australe genome is composed of various repeat sequences, differing from those of G. arboreum (85.39%), G. hirsutum (69.86%) and G. barbadense (69.83%). The G. australe genome showed closer collinear relationships with the genome of G. arboreum than G. raimondii and has undergone less extensive genome reorganization than the G. arboreum genome. Selection signature and transcriptomics analyses implicated multiple genes in disease resistance responses, including GauCCD7 and GauCBP1, and experiments revealed induction of both genes by Verticillium dahliae and by the plant hormones strigolactone (GR24), salicylic acid (SA) and methyl jasmonate (MeJA). Experiments using a Verticillium-resistant domesticated G. barbadense cultivar confirmed that knockdown of the homologues of these genes caused a significant reduction in resistance against Verticillium dahliae. Moreover, knockdown of a newly identified gland-associated gene GauGRAS1 caused a glandless phenotype in partial tissues using G. australe. The G. australe genome represents a valuable resource for cotton research and distant relative breeding as well as for understanding the evolutionary history of crop genomes.

Transcriptome Analysis Provides Insights into Grain Filling in Foxtail Millet (Setaria italica L.).

Grain filling is an importantly developmental process which is associated with the yield and quality of foxtail millet (Setaria italic L.). However, the molecular mechanisms of grain filling are rarely reported in foxtail millet. In our study, RNA-seq was performed to investigate the transcriptional dynamics and identify the key genes involved in grain filling in foxtail millet at five different developmental stages. A total of 11,399 differentially expressed genes (DEGs), including 902 transcription factors (TFs), were identified. Certain important genes involved in grain filling were discovered through a function annotation and temporal expression patterns analysis. These genes included genes associated with starch biosynthesis, cell-wall invertases, hormone signal transduction, and polyamine metabolism pathways. The expression levels of seven randomly selected DEGs were validated by a quantitative real-time polymerase chain reaction (qRT-PCR). This study provides the first insight into the changes in the gene expression of grain filling at different developmental stages in foxtail millet. These results could help understand the complex molecular mechanisms of the panicle formation in foxtail millet and other cereal crops.

Genome-wide analysis of the cotton G-coupled receptor proteins (GPCR) and functional analysis of GTOM1, a novel cotton GPCR gene under drought and cold stress.

BACKGROUND: The efficient detection and initiation of appropriate response to abiotic stresses are important to plants survival. The plant G-protein coupled receptors (GPCRs) are diverse membranous proteins that are responsible for signal transduction. RESULTS: In this research work, we identified a novel gene of the GPCR domain, transformed and carried out the functional analysis in Arabidopsis under drought and cold stresses. The transgenic lines exposed to drought and cold stress conditions showed higher germination rate, increased root length and higher fresh biomass accumulation. Besides, the levels of antioxidant enzymes, glutathione (GSH) and ascorbate peroxidase (APX) exhibited continuously increasing trends, with approximately threefold higher than the control, implying that these ROS-scavenging enzymes were responsible for the detoxification of ROS induced by drought and cold stresses. Similarly, the transgenic lines exhibited stable cell membrane stability (CMS), reduced water loss rate in the detached leaves and significant values for the saturated leaves compared to the wild types. Highly stress-responsive miRNAs were found to be targeted by the novel gene and based on GO analysis; the protein encoded by the gene was responsible for maintaining an integral component of membrane. In cotton, the virus-induced gene silencing (VIGS) plants exhibited a higher susceptibility to drought and cold stresses compared to the wild types. CONCLUSION: The novel GPCR gene enhanced drought and cold stress tolerance in transgenic Arabidopsis plants by promoting root growth and induction of ROS scavenging enzymes. The outcome showed that the gene had a role in enhancing drought and cold stress tolerance, and can be further exploited in breeding for more stress-resilient and tolerant crops.

Transcriptomic and biochemical analysis of upland cotton (Gossypium hirsutum) and a chromosome segment substitution line from G. hirsutum × G. barbadense in response to Verticillium dahliae infection.

BACKGROUND: Verticillium wilt (VW), also known as "cotton cancer," is one of the most destructive diseases in global cotton production that seriously impacts fiber yield and quality. Despite numerous attempts, little significant progress has been made in improving the VW resistance of upland cotton. The development of chromosome segment substitution lines (CSSLs) from Gossypium hirsutum × G. barbadense has emerged as a means of simultaneously developing new cotton varieties with high-yield, superior fiber, and resistance to VW. RESULTS: In this study, VW-resistant investigations were first conducted in an artificial greenhouse, a natural field, and diseased nursery conditions, resulting in the identification of one stably VW-resistant CSSL, MBI8255, and one VW-susceptible G. hirsutum, CCRI36, which were subsequently subjected to biochemical tests and transcriptome sequencing during V991 infection (0, 1, and 2 days after inoculation). Eighteen root samples with three replications were collected to perform multiple comparisons of enzyme activity and biochemical substance contents. The findings indicated that VW resistance was positively correlated with peroxidase and polyphenol oxidase activity, but negatively correlated with malondialdehyde content. Additionally, RNA sequencing was used for the same root samples, resulting in a total of 77,412 genes, of which 23,180 differentially expressed genes were identified from multiple comparisons between samples. After Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis on the expression profiles identified using Short Time-series Expression Miner, we found that the metabolic process in the biological process, as well as the pathways of phenylpropanoid biosynthesis and plant hormone signal transduction, participated significantly in the response to VW. Gene functional annotation and expression quantity analysis indicated the important roles of the phenylpropanoid metabolic pathway and oxidation-reduction process in response to VW, which also provided plenty of candidate genes related to plant resistance. CONCLUSIONS: This study concentrates on the preliminary response to V991 infection by comparing the VW-resistant CSSL and its VW-susceptible recurrent parent. Not only do our findings facilitate the culturing of new resistant varieties with high yield and superior performance, but they also broaden our understanding of the mechanisms of cotton resistance to VW.

Functional characterization of Gh_A08G1120 (GH3.5) gene reveal their significant role in enhancing drought and salt stress tolerance in cotton.

BACKGROUND: Auxins play an important role in plant growth and development; the auxins responsive gene; auxin/indole-3-acetic acid (Aux/IAA), small auxin-up RNAs (SAUR) and Gretchen Hagen3 (GH3) control their mechanisms. The GH3 genes function in homeostasis by the catalytic activities in auxin conjugation and bounding free indole-3-acetic acid (IAA) to amino acids. RESULTS: In our study, we identified the GH3 genes in three cotton species; Gossypium hirsutum, Gossypium arboreum and Gossypium raimondii, analyzed their chromosomal distribution, phylogenetic relationships, cis-regulatory element function and performed virus induced gene silencing of the novel Gh_A08G1120 (GH3.5) gene. The phylogenetic tree showed four clusters of genes with clade 1, 3 and 4 having mainly members of the GH3 of the cotton species while clade 2 was mainly members belonging to Arabidopsis. There were no paralogous genes, and few orthologous genes were observed between Gossypium and other species. All the GO terms were detected, but only 14 genes were found to have described GO terms in upland cotton, more biological functions were detected, as compared to the other functions. The GH3.17 subfamily harbored the highest number of the cis-regulatory elements, most having promoters towards dehydration-responsiveness. The RNA expression analysis revealed that 10 and 8 genes in drought and salinity stress conditions respectively were upregulated in G. hirsutum. All the genes that were upregulated in plants under salt stress conditions were also upregulated in drought stress; moreover, Gh_A08G1120 (GH3.5) exhibited a significant upregulation across the two stress factors. Functional characterization of Gh_A08G1120 (GH3.5) through virus-induced gene silencing (VIGS) revealed that the VIGS plants ability to tolerate drought and salt stresses was significantly reduced compared to the wild types. The chlorophyll content, relative leaf water content (RLWC), and superoxide dismutase (SOD) concentration level were reduced significantly while malondialdehyde concentration and ion leakage as a measure of cell membrane stability (CMS) increased in VIGS plants under drought and salt stress conditions. CONCLUSION: This study revealed the significance of the GH3 genes in enabling the plant's adaptation to drought and salt stress conditions as evidenced by the VIGS results and RT-qPCR analysis.

Microdissection of the Ah01 chromosome in upland cotton and microcloning of resistance gene anologs from the single chromosome.

BACKGROUND: Chromosome microdissection is one of the most important techniques in molecular cytogenetic research. Cotton (Gossypium Linnaeus, 1753) is the main natural fiber crop in the world. The resistance gene analog (RGA) cloning after its single chromosome microdissection can greatly promote cotton genome research and breeding. RESULTS: Using the linker adaptor PCR (LA-PCR) with the primers of rice disease-resistance homologues, three nucleotide sequences PS016 (KU051681), PS054 (KU051682), and PS157 (KU051680) were obtained from the chromosome Ah01 of upland cotton (cv. TM-1). The Blast results showed that the three sequences are the nucleotide binding site-leucine rich repeat (NBS-LRR) type RGAs. Clustering results indicated that they are homologous to these published RGAs. Thus, the three RGAs can definitely be confirmed as NBS-LRR class of RGAs in upland cotton. CONCLUSIONS: Using single chromosome microdissection technique, DNA libraries containing cotton RGAs were obtained. This technique can promote cotton gene cloning, marker development and even the improvement of cotton genome research and breeding.

Exploring the plasmodesmata callose-binding protein gene family in upland cotton: unraveling insights for enhancing fiber length.

Plasmodesmata are transmembrane channels embedded within the cell wall that can facilitate the intercellular communication in plants. Plasmodesmata callose-binding (PDCB) protein that associates with the plasmodesmata contributes to cell wall extension. Given that the elongation of cotton fiber cells correlates with the dynamics of the cell wall, this protein can be related to the cotton fiber elongation. This study sought to identify PDCB family members within the Gossypium. hirsutum genome and to elucidate their expression profiles. A total of 45 distinct family members were observed through the identification and screening processes. The analysis of their physicochemical properties revealed the similarity in the amino acid composition and molecular weight across most members. The phylogenetic analysis facilitated the construction of an evolutionary tree, categorizing these members into five groups mainly distributed on 20 chromosomes. The fine mapping results facilitated a tissue-specific examination of group V, revealing that the expression level of GhPDCB9 peaked five days after flowering. The VIGS experiments resulted in a marked decrease in the gene expression level and a significant reduction in the mature fiber length, averaging a shortening of 1.43-4.77 mm. The results indicated that GhPDCB9 played a pivotal role in the cotton fiber development and served as a candidate for enhancing cotton yield.

Widespread incomplete lineage sorting and introgression shaped adaptive radiation in the Gossypium genus.

Cotton (Gossypium) stands as a crucial economic crop, serving as the primary source of natural fiber for the textile sector. However, the evolutionary mechanisms driving speciation within the Gossypium genus remain unresolved. In this investigation, we leveraged 25 Gossypium genomes and introduced four novel assemblies-G. harknessii, G. gossypioides, G. trilobum, and G. klotzschianum (Gklo)-to delve into the speciation history of this genus. Notably, we encountered intricate phylogenies potentially stemming from introgression. These complexities are further compounded by incomplete lineage sorting (ILS), a factor likely to have been instrumental in shaping the swift diversification of cotton. Our focus subsequently shifted to the rapid radiation episode during a concise period in Gossypium evolution. For a recently diverged lineage comprising G. davidsonii, Gklo, and G. raimondii, we constructed a finely detailed ILS map. Intriguingly, this analysis revealed the non-random distribution of ILS regions across the reference Gklo genome. Moreover, we identified signs of robust natural selection influencing specific ILS regions. Noteworthy variations pertaining to speciation emerged between the closely related sister species Gklo and G. davidsonii. Approximately 15.74% of speciation structural variation genes and 12.04% of speciation-associated genes were estimated to intersect with ILS signatures. These findings enrich our understanding of the role of ILS in adaptive radiation, shedding fresh light on the intricate speciation history of the Gossypium genus.

Genome-wide identification and functional analysis of the SiCIN gene family in foxtail millet (Setaria italica L.).

Cell wall invertase (CIN) is a vital member of plant invertase (INV) and plays a key role in the breakdown of sucrose. This enzyme facilitates the hydrolysis of sucrose into glucose and fructose, which is crucial for various aspects of plant growth and development. However, the function of CIN genes in foxtail millet (Setaria italica) is less studied. In this research, we used the blast-p of NCBI and TBtools for bidirectional comparison, and a total of 13 CIN genes (named SiCINs) were identified from foxtail millet by using Arabidopsis and rice CIN sequences as reference sequences. The phylogenetic tree analysis revealed that the CIN genes can be categorized into three subfamilies: group 1, group 2, and group 3. Furthermore, upon conducting chromosomal localization analysis, it was observed that the 13 SiCINs were distributed unevenly across five chromosomes. Cis-acting elements of SiCIN genes can be classified into three categories: plant growth and development, stress response, and hormone response. The largest number of cis-acting elements were those related to light response (G-box) and the cis-acting elements related to seed-specific regulation (RY-element). qRT-PCR analysis further confirmed that the expression of SiCIN7 and SiCIN8 in the grain was higher than that in any other tissues. The overexpression of SiCIN7 in Arabidopsis improved the grain size and thousand-grain weight, suggesting that SiCIN7 could positively regulate grain development. Our findings will help to further understand the grain-filling mechanism of SiCIN and elucidate the biological mechanism underlying the grain development of SiCIN.