Drought stress significantly affects the growth, development, and yield of cotton, triggering the response of multiple genes. Among them, ascorbate peroxidase (APX) is one of the important antioxidant enzymes in the metabolism of reactive oxygen species in plants, and APX enhances the ability of plants to resist oxidation, thus increasing plant stress tolerance. Therefore, enhancing the activity of APX in cells is crucial to improving plant stress resistance. Previous studies have isolated differentially expressed proteins under drought stress (GhAPX7) in drought-resistant (KK1543) and drought-sensitive (XLZ26) plants. Thus, this study analyzed the expression patterns of GhAPX7 in different cotton tissues to verify the drought resistance function of GhAPX7 and explore its regulatory pathways. GhAPX7 had the highest expression in cotton leaves, which significantly increased under drought stress, suggesting that GhAPX7 is essential for improving antioxidant capacity and enzyme activities in cotton. GhAPX7 silencing indirectly affects pronounced leaf yellowing and wilting in drought-resistant and drought-sensitive plants under drought stress. Malondialdehyde (MDA) content was significantly increased and chlorophyll and proline content and APX enzyme activity were generally decreased in silenced plants compared to the control. This result indicates that GhAPX7 may improve drought resistance by influencing the contents of MDA, chlorophyll, proline, and APX enzyme activity through increased expression levels. Transcriptome analysis revealed that the drought-related differentially expressed genes between the control and treated groups enriched plant hormone signal transduction, MAPK signaling, and plant–pathogen interaction pathways. Therefore, the decreased expression of GhAPX7 significantly affects the expression levels of genes in these three pathways, reducing drought resistance in plants. This study provides insights into the molecular mechanisms of GhAPX7 and its role in drought resistance and lays a foundation for further research on the molecular mechanisms of response to drought stress in cotton.

Fuzzless Gossypium hirsutum mutants are ideal materials for investigating cotton fiber initiation and development. In this study, we used the fuzzless G. hirsutum mutant Xinluzao 50 FLM as the research material and combined it with other fuzzless materials for verification by RNA sequencing to explore the gene expression patterns and differences between genes in upland cotton during the fuzz period. A gene ontology (GO) enrichment analysis showed that differentially expressed genes (DEGs) were mainly enriched in the metabolic process, microtubule binding, and other pathways. A weighted gene co-expression network analysis (WGCNA) showed that two modules of Xinluzao 50 and Xinluzao 50 FLM and four modules of CSS386 and Sicala V-2 were highly correlated with fuzz. We selected the hub gene with the highest KME value among the six modules and constructed an interaction network. In addition, we selected some genes with high KME values from the six modules that were highly associated with fuzz in the four materials and found 19 common differential genes produced by the four materials. These 19 genes are likely involved in the formation of fuzz in upland cotton. Several hub genes belong to the arabinogalactan protein and GDSL lipase, which play important roles in fiber development. According to the differences in expression level, 4 genes were selected from the 19 genes and tested for their expression level in some fuzzless materials. The modules, hub genes, and common genes identified in this study can provide new insights into the formation of fiber and fuzz, and provide a reference for molecular design breeding for the genetic improvement of cotton fiber.

[目的]评价陆地棉种质资源材料的耐热性及筛选指标,为选育耐热性棉花品种及研究耐热性机理提供参考依据.[方法]基于2 年试验,选用24 份棉花种质资源,随机区组设计,测定相关指标,综合评价 24 份陆地棉种质资源材料的耐热性.[结果]24 份陆地棉资源材料可分为3 类.其中第一类为耐热材料,第二类为中间型材料,第三类为敏热型材料,建立陆地棉耐热性的评价模型:D =-0.111+0.175X2+0.470X3+1.211X4(R2 =0.9148),建立的最优回归方程可以预测陆地棉材料的耐热性.籽棉产量、单铃重和皮棉产量可作为棉花大田耐热的评价指标,而株高和衣分与耐热性无关.[结论]筛选出4 份耐热陆地棉资源材料,建立了以籽棉产量、单铃重和皮棉产量作为陆地棉大田耐热性的评价指标,以过氧化物酶、丙二醛和叶绿素含量作为室内耐热性的评价指标.

Background Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is the central enzyme of glycolysis and plays important regulatory roles in plant growth and development and responses to adverse stress conditions. However, studies on the characteristics and functions of cotton GAPDH family genes are still lacking. Methods In this study, genome-wide identification of the cotton GAPDH gene family was performed, and the phylogeny, gene structures, promoter progenitors and expression profiles of upland cotton GAPDH gene family members were explored by bioinformatics analysis to highlight potential functions. The functions of GhGAPDH9 in response to drought stress were initially validated based on RNA-seq, qRT‒PCR, VIGS techniques and overexpression laying a foundation for further studies on the functions of GAPDH genes. Results This study is the first systematic analysis of the cotton GAPDH gene family, which contains a total of 84 GAPDH genes, among which upland cotton contains 27 members. Quantitative, phylogenetic and covariance analyses of the genes revealed that the GAPDH gene family has been conserved during the evolution of cotton. Promoter analysis revealed that most cis-acting elements were related to MeJA and ABA. Based on the identified promoter cis-acting elements and RNA-seq data, it was hypothesized that Gh_GAPDH9, Gh_GAPDH11, Gh_GAPDH19 and Gh_GAPDH21 are involved in the response of cotton to abiotic stress. The expression levels of the Gh_GAPDH9 gene in two drought-resistant and two drought-sensitive materials were analyzed by qRT‒PCR and found to be high early in the treatment period in the drought-resistant material. The silencing of Gh_GAPDH9 based on virus-induced gene silencing (VIGS) technology resulted in significant leaf wilting or whole-plant dieback in silenced plants after drought stress compared to the control. The content of—malondialdehyde (MDA) in cotton leaves was significantly increased, and the content of proline (Pro) and chlorophyll (Chl) was reduced. In addition, the leaf wilting and dryness of transgenic lines under drought stress were lower than those of wild-type Arabidopsis, indicating that Gh_GAPDH9 is a positive regulator of drought resistance. In conclusion, our results demonstrate that GAPDH genes play an important role in the response of cotton to abiotic stresses and provide preliminary validation of the function of the Gh_GAPDH9 gene under drought stress. These findings provide an important theoretical basis for further studies on the function of the Gh_GAPDH9 gene and the molecular mechanism of the drought response in cotton.

Gossypium barbadense possesses a superior fiber quality because of its fiber length and strength. An in-depth analysis of the underlying genetic mechanism could aid in filling the gap in research regarding fiber strength and could provide helpful information for Gossypium barbadense breeding. Three quantitative trait loci related to fiber strength were identified from a Gossypium barbadense recombinant inbred line (PimaS-7 x 5917) for further analysis. RNA sequencing was performed in the fiber tissues of PimaS-7 x 5917 0-35 days postanthesis. Four specific modules closely related to the secondary wall-thickening stage were obtained using the weighted gene coexpression network analysis. In total, 55 genes were identified as differentially expressed from 4 specific modules. Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes were used for enrichment analysis, and Gbar_D11G032910, Gbar_D08G020540, Gbar_D08G013370, Gbar_D11G033670, and Gbar_D11G029020 were found to regulate fiber strength by playing a role in the composition of structural constituents of cytoskeleton and microtubules during fiber development. Quantitative real-time PCR results confirmed the accuracy of the transcriptome data. This study provides a quick strategy for exploring candidate genes and provides new insights for improving fiber strength in cotton.

植物生长素转运蛋白之一的PIN针状蛋白(PIN-formed acicular protein,PIN)影响棉花(Gossypium spp.)纤维的品质.研究海岛棉(G.barbadense)GbPIN1a(GenBank No.KAB2059844.1)基因在棉纤维中的表达特征,可为今后探究棉花纤维品质发育机理提供参考.本研究借助PCR技术克隆该基因,进行生物信息学分析,构建了含GFP的瞬时表达载体,进行细胞定位分析.研究发现GbPIN1a基因编码区全长1758 bp,编码585个氨基酸,氨基酸序列包含2个Mem trans保守结构域;GbPIN1a基因在纤维发育的15、20 DPA(开花后天数,days post anthesis)呈现高表达趋势且与同时期的生长素含量呈显著负相关(P<0.05);在根和茎中的表达量显著高于叶(P<0.05),存在组织特异性;在受到外源植物生长素刺激的情况下表达量会出现波动;表达产物被定位于细胞膜上.本研究结果初步表明GbPIN1a基因可能参与调控棉花的纤维发育,为今后研究PIN1a基因提供基础资料.

[目的]运用SSR(Simple sequence repeat)分子标记关联分析204份海岛棉种质资源的抗枯萎病性状,为抗枯萎病棉花育种材料的利用、分子标记辅助选择提供参考.[方法]采用前期实验室研究的抗病基因序列,设计出40对SSR分子标记,采用聚丙烯酰胺凝胶电泳检测筛选标记引物特异性,并鉴定204份海岛棉种质资源.[结果](1)从40对引物中筛选出6对引物具有多态性,多次重复后选用CHS-04和CHS-05两个分子标记,均来自类黄酮代谢途径;(2)引物CHS-04扩增条带与群体抗病性一致率介于41.67％～52.94％,平均值为46.67％;引物CHS-05扩增条带与群体材料一致率介于30.39％～54.68％,平均值为35.78％.[结论]引物CHS-04和CHS-05可以作为海岛棉枯萎病抗性的分子标记鉴定指标.

Background PIN proteins are an important class of auxin polar transport proteins that play an important regulatory role in plant growth and development. However, their characteristics and functions have not been identified in Gossypium barbadense. Methods PIN family genes were identified in the cotton species G. barbadense, Gossypium hirsutum, Gossypium raimondii, and Gossypium arboreum, and detailed bioinformatics analyses were conducted to explore the roles of these genes in G. barbadense using transcriptome data and quantitative reverse-transcription polymerase chain reaction (qRT-PCR) technology. Functional verification of the genes was performed using virus-induced gene silencing (VIGS) technology. Results A total of 138 PIN family genes were identified in the four cotton species; the genes were divided into seven subgroups. GbPIN gene family members were widely distributed on 20 different chromosomes, and most had repeated duplication events. Transcriptome analysis showed that some genes had differential expression patterns in different stages of fiber development. According to ‘PimaS-7’ and ‘5917’ transcript component association analysis, the transcription of five genes was directly related to endogenous auxin content in cotton fibers. qRT-PCR analysis showed that the GbPIN7 gene was routinely expressed during fiber development, and there were significant differences among materials. Transient silencing of the GbPIN7 gene by VIGS led to significantly higher cotton plant growth rates and significantly lower endogenous auxin content in leaves and stems. This study provides comprehensive analyses of the roles of PIN family genes in G. barbadense and their expression during cotton fiber development. Our results will form a basis for further PIN auxin transporter research.

Background: As a ubiquitous acid-regulating protein family in eukaryotes, 14-3-3 proteins are widely involved in the growth and development of plants. With the development of the third-generation sequencing technology and the smooth completion of the cotton genome work, it is possible to explore the existence and distribution of the 14-3-3 protein family in cotton.Results: In this paper, 33, 33, 17 and 18 members were identified from this family in Gossypium hirsutum((AD)1), G. barbadense((AD)2), G. arboreum(A2) and G. raimondii(D5), respectively. In particular, evolution analysis, structure analysis and functional expression analysis of this protein family in G. hirsutum((AD)1) were carried out. The results showed that compared with Arabidopsis and rice, the phylogenetic tree and gene structure clearly divided the 14-3-3 protein family into two subgroups in G. hirsutum((AD)1), the ε group and the non-ε group; Analysis of transcriptome expression patterns revealed that this family was significantly induced to express under abiotic stress; Most 14-3-3 proteins have a large number of cis-acting elements related to growth, development and abiotic stress in the promoter region, among which elements related to drought stress account for the largest proportion; The results of qRT-PCR showed that the expression of 14-3-3 protein had significant differences under drought stress.Conclusions: In summary, this study signified that the 14-3-3 protein family is relatively conserved in the evolutionary expansion of cotton, and may be involved in the growth and development of plants and the mechanism of stress resistance. These results provide an important theoretical and experimental basis for further analysis and verification of the function of 14-3-3 protein in cotton.

Background Domain of unknown function 668 (DUF668) may play a crucial role in the plant growth and developmental response to adverse stress. However, our knowledge of the function of the DUF668 gene family is limited. Results Our study was conducted based on the DUF668 gene family identified from cotton genome sequencing. Phylogenetic analysis showed that the DUF668 family genes can be classified into four subgroups in cotton. We identified 32 DUF668 genes, which are distributed on 17 chromosomes and most of them located in the nucleus of Gossypium hirsutum. Gene structure and motif analyses revealed that the members of the DUF668 gene family can be clustered in G. hirsutum into two broad groups, which are relatively evolutionarily conserved. Transcriptome data analysis showed that the GhDUF668 genes are differentially expressed in different tissues under various stresses (cold, heat, drought, salt, and Verticillium dahliae ), and expression is generally increased in roots and stems. Promoter and expression analyses indicated that Gh_DUF668–05, Gh_DUF668–08, Gh_DUF668–11, Gh_DUF668–23 and Gh_DUF668–28 in G. hirsutum might have evolved resistance to adverse stress. Additionally, qRT-PCR revealed that these 5 genes in four cotton lines, KK1543 (drought resistant), Xinluzao 26 (drought sensitive), Zhongzhimian 2 (disease resistant) and Simian 3 (susceptible), under drought and Verticillium wilt stress were all significantly induced. Roots had the highest expression of these 5 genes before and after the treatment. Among them, the expression levels of Gh_DUF668–08 and Gh_DUF668–23 increased sharply at 6 h and reached a maximum at 12 h under biotic and abiotic stress, which showed that they might be involved in the process of adverse stress resistance in cotton. Conclusion The significant changes in GhDUF668 expression in the roots after adverse stress indicate that GhDUF668 is likely to increase plant resistance to stress. This study provides an important theoretical basis for further research on the function of the DUF668 gene family and the molecular mechanism of adverse stress resistance in cotton.