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15 July 2020, Volume 32 Issue 4
    Genome-Wide Identification and Analysis of Dof Gene Family in Upland Cotton
    Ju Longzhen, Zhao Ting, Fang Lei, Hu Yan, Zhang Tianzhen
    2020, 32(4):  279-291.  doi:10.11963/1002-7807.jlzhy.20200630
    Abstract ( 500 )   PDF (14548KB) ( 200 )  
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    [Objective] The DNA binding with one finger proteins (Dof) are plant-specific transcription factors that play a very important role in plant growth, development and in response to abiotic stress. The aim of this study was to provide a reference for further functional study of the Dof genes associated with plant growth and development by genome-wide analysis of Dof transcription factor family in Gossypium hirsutum. [Method] Using the recently released upland cotton genomic data, the Dof genes in G. hirsutum were identified by bioinformatic method. The physico-chemical properties, sequence characteristics, gene duplication, phylogeny and expression pattern of Dof genes were analyzed. [Result] A total of 118 Dof genes were identified in G. hirsutum. Based on phylogenetic analysis, these Dof genes were divided into nine subfamilies, and the genes within the same subfamily had similar exon-intron organization and motif distribution. Gene duplication analysis revealed that whole genome duplication was the main contributor to the expansion of Dof genes in G. hirsutum. Cis-acting element analysis showed the promoters of Dof genes had cis-acting regulatory elements involved in stresses responses to different plant hormones. The results of RNA-seq data analysis revealed that Dof genes expression in G. hirsutum was classified into three types which had different expression pattern in different tissues, developmental stages and stresses, indicating gene functional differentiation. [Conclusion] The genome-wide identification and analysis of Dof gene family in G. hirsutum is helpful to understand the evolution and function, and provides a support for future studies.
    Cloning and Expression Profiling of Cuticular Protein Genes of Spodoptera litura
    Zhao Peng, Zhang Shuai, Zhao Chenchen, Hu Fangmei, Cui Jinjie, Li Shaoqin
    2020, 32(4):  292-304.  doi:10.11963/1002-7807.zplsq.20200521
    Abstract ( 169 )   PDF (5483KB) ( 42 )  
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    [Objective] The expression pattern of fourteen cuticular protein (CP) genes (ICPG) in Spodoptera litura were analyzed in this study, which provided reference for further study on the function of cuticular proteins in the growth and development of S. litura. [Method] Polymerase chain reaction (PCR) was used to amplify the cDNAs of the genes encoding CPs of S. litura. Their nucleotide sequences and corresponding amino acid sequences were analyzed, and lepidopteran phylogenetic tree was constructed using MEGA5.2 software. The expression of CP genes was analyzed in the third to the sixth instar larvae and 2-day-old adult genital organs by the reverse transcription PCR (RT-PCR) technique. [Result] Fourteen cDNAs coding 14 CPs of S. litura were obtained. The sequence analysis showed 14 CPs harbor the conserved motif of 35-36 amino acids that is a typical feature of chitin-binding proteins and share high sequence homology with CPs of Helicoverpa armigera. RT-PCR results showed that these CP genes were expressed in the third to the sixth instar larvae and 2-day-old adults. In particular, their relative expression levels are higher in the cuticulars of older larvae and lower in the fat bodies of younger larvae. In addition, the relative expression of 14 CP genes was low in testes and ovaries. The highest relative expression of SlICPG-11 and SlICPG-14 in the pupal stage. [Conclusion] In this study, the cDNA of 14 S. litura CP genes were cloned. Their expression exhibited tissue and developmental stage specificity indicating their putative roles in S. litura development. It lays the foundation for target site selection for the control of S. litura.
    Cloning and Functional Analysis of Low Temperature Response Gene, GhZAT10, in  Upland Cotton at Seedling Stage
    Yang Yongfei , Ge Changwei, Shen Qian, Zhang Siping, Liu Shaodong, Ma Huijuan, Chen Jing, Liu Ruihua, Li Shicong, Zhao Xinhua, Li Zhikun, Pang Chaoyou
    2020, 32(4):  305-315.  doi:10.11963/1002-7807.yyfpcy.20200703
    Abstract ( 162 )   PDF (6553KB) ( 67 )  
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    [Objective] The aim of this study is to identify chilling tolerant genes which can provide a basis for breeding upland cotton cultivars with tolerance to chilling. [Method] The GhZAT10 (Zinc finger of Arabidopsis thaliana 10) gene was cloned, and its expression in roots, stems, and leaves was analyzed with cold treatment by quantitative real-time polymerase chain reaction (qRT-PCR). Then we analyzed the structural characteristics of GhZAT10, protein properties and constructed the phylogenetic tree by bioinformatics methods. Next, we constructed subcellular localization vector 35S::GhZAT10-GFP by Gateway technology. Finally, we studied the effect of GhZAT10 in chilling response by virus induced gene silencing of cotton. [Result] The Open reading frame(ORF) length of GhZAT10 gene is 813 bp (base pair), encoding 270 amino acid residues. The expression of GhZAT10 in roots was higher than that in stems and leaves, and up-regulated after chilling stress in these tissues. GhZAT10 does not possess signal peptides or transmembrane helices; its activity is closely related to the phosphorylation regulation. GhZAT10 has homology with the ZAT10 of Theobroma cacao, Arabidopsis thaliana and Citrus sinensis. GhZAT10 protein is located in the nucleus. The GhZAT10-silenced cotton plants were more sensitive to low temperature than wild type. [Conclusion] GhZAT10 belongs to C2H2 zinc finger protein, and plays a positive role to response the chilling stress in upland cotton.
    Genome-Wide Identification and Functional Analysis of the NF-YA Gene Family in Gossypium hirsutum L.
    Pan Ao, Wang Jingjing, Sun Fulai, Zhang Jingxia, Gao Yang, Du Zhaohai, Jiao Mengjia, Zhang Jun, Wang Furong, Liu Zhi
    2020, 32(4):  316-328.  doi:10.11963/1002-7807.palz.20200611
    Abstract ( 178 )   PDF (7726KB) ( 74 )  
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    [Objective] The NF-YA gene family was identified in upland cotton (Gossypium hirsutum L.), and their expression characteristics were analyzed to discover the genes related to cotton flowering regulation. [Method] Using bioinformatic methods, the NF-YA genes were identified and their physical and chemical properties, gene structure, cis-element, collinearity, Ka/K swere investigated. The expression pattern was performed using quantitative real-time polymerase chain reaction (qRT-PCR). The gene function was verified by virus-induced gene silencing (VIGS). [Result] A total of 29 GhNF-YA genes were identified from upland cotton genome, these genes were divided into five clades and distributed on 18 chromosomes. The expansion of the NF-YA genes in upland cotton was mainly driven by whole genome duplication and segmental duplication. A lot of light responding elements were identified in the promoter regions of GhNF-YA genes. According to the expression profiling, the GhNF-YA genes were highly expressed in stems and leaves. Meanwhile, analysis of qRT-PCR showed that 12 GhNF-YA genes had a relatively high expression in the leaf of both early-maturing cotton cultivar Lumianyan 19 and late-maturing cotton cultivar Lumianyan 37 from three- to six-leaf stage, and expression of the most genes was significantly different between the two cultivars. The function of GhNF-YA18 in Lumianyan 37 was verified by VIGS, and the results indicated that the bud emergence in silenced plants was 11 d earlier than that of unsilenced control plants. In addition, the expression of GhNF-YA18 was lower in silenced plants than unsilenced plants at albino and budding stages. [Conclusion] In this study, we identified and characterized the 29 GhNF-YA genes in upland cotton. The results will provide a foundation for future studies in molecular regulatory mechanisms of cotton early fiowering.
    Biocontrol Effect of Bacillus velezensis Strain SZAD1 on Verticillium dahliae
    Zhang Qiong, Zabihullah Sherzad, Tang Canming
    2020, 32(4):  329-338.  doi:10.11963/1002-7807.zqtcm.20200609
    Abstract ( 159 )   PDF (5811KB) ( 37 )  
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    [Objective] Verticillium wilt is the main fungal disease of cotton, lacking high resistant varieties and safe and effective chemical fungicides. Biological control of Verticillium wilt is of great significance to cotton production. The purpose of this study was to analyze the antagonistic activity and biocontrol potential of endophytic bacteria in cotton against Verticillium dahliae. [Method] SZAD1 was identified by 16S rDNA and gyrB sequence analysis. The ability of SZAD1 to inhibit the mycelial growth of V.dahliae strain VD080 was tested by the agar plate test, and the control effect on Verticillium wilt was tested by the seed soaking and root filling method. The agar plate containing carboxy methyl cellulose (CMC) or chitin was prepared to test whether the strain can secrete cellulase and chitinase, and the enzyme activity was determined by the dinitrosalicylic acid reagent method. [Result] SZAD1 belongs to Bacillus velezensis. SZAD1 can significantly inhibit the growth of VD080 mycelium. The control effect of SZAD1 on Verticillium wilt was 60.10% and 56.00% in the seed soaking and root filling  experiments respectively. SZAD1 strain can produce cellulase and chitinase with the maximum activity at 72 h after culture. The antibacterial effect of fermented supernatant was the strongest at 72 h after culture. The content of VD080 spores in liquid potato dextrose agar medium decreased by 30.20% and 96.53% compared with the control respectively after adding supernatant with concentrations of 25 mg·L-1 and 50 mg·L-1. [Conclusion] The supernatant of bacteria SZAD1 can reduce the colonization of VD080 in the stem of cotton plant by root filling and decreased the withering degree of cotton leaves. SZAD1 has the potential of controlling cotton Verticillium wilt as a biocontrol bacteria.
    The Effects of Increased-Density on Canopy Apparent Photosynthesis, Dry Matter Accumulation and Distribution of Cotton under Late-Sown Condition
    Li Hui, Wan Hualong, Tian Liwen, Liu Liantao, Zhang Yongjiang, Bai Zhiying, Zhang Ke, Wang Guoping, Sun Hongchun, Li Cundong
    2020, 32(4):  339-347.  doi:10.11963/1002-7807.lhlcd.20200622
    Abstract ( 141 )   PDF (3579KB) ( 75 )  
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    [Objective] The effects of high planting densities on canopy apparent photosynthesis, leaf area index, dry matter accumulation and distribution features of cotton were studied, with purpose to elucidate the suitable density for late cotton-sown system in the Yellow River cultivation region. [Method] Field experiments including two planting density treatments (D1 of 9.0×104 plant·hm-2 and D2 of 12.0×104 plant·hm-2) were conducted during the 2017 and 2018 growth seasons, using cultivars Nongda 601(ND-601) and Guoxin Cotton 9(GX-9) in 2017 and Nongda 601 in 2018 as the materials. The effects of densities on photosynthetic characteristics and yield components were investigated. [Result] At fast growth stage, the leaf area index under D2 was significantly higher than that of D1, which sustained longer peak duration under the former condition. These results suggested that higher density can promote the generation of enlarged canopy structure at vigorous growth stage. However, much intensified leaf area index resulted in population shading and reduced canopy apparent photosynthesis. Although D2 treatment was shown to benefit the biomass accumulation of the population plants and the vegetative organs. But the enhanced vegetative growth led to restriction on the reproductive tissue, which resulted in lowered nutrient distribution to reproductive organ; the increase of density was no significant difference on boll weight and lint percentage. Perhaps because of the interannual climate variation, the results of other yield components obtained across the two growth seasons were inconsistent with each other. [Conclusion] Under the suitable conditions, D1 treatment is beneficial to establish the reasonable population structure and to achieve stable yield. In contrast, the D2 treatment has the potential to achieve higher yield. Our results suggested that D2 together with adoption of chemical control technology can help high-yielding cultivation, through increasing population dry matter amount, improving nutrient distribution across organs, and elevating harvest index of the cotton plants.
    Identification and Analysis of Fiber Quality and Yield Related Traits of Interspecific  (Gossypium hirsutum L. × G. barbadense L.) Hybrids
    Li Tengyu, Xu Chao, Li Yaoming, Gou Chengfei, Hong Zhu, Ding Mingquan, Sun Chendong
    2020, 32(4):  348-359.  doi:10.11963/1002-7807.ltyscd.20200612
    Abstract ( 158 )   PDF (1238KB) ( 48 )  
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    [Objective] The aim of this study is to study the hereditary of heterosis of fiber quality and yield-related traits in the upland-island interspecific hybrids, and breed new interspecific hybrid varieties with high yield and fine fiber quality. [Method] In this study, 12 upland cotton materials and 5 sea-island cotton materials were selected to determine the fiber quality and yield traits of their parents and F1 in Lin’an, Zhejiang and Sanya, Hainan. [Result] It was found that fiber length and fiber strength of F1 (Gossypium hirsutum × G. barbadense) generally had significant mid-parent heterosis (MPH), some hybrid combinations showed strong over-parent heterosis (OPH), fiber length had a small coefficient of variation between the two places and could be stably inherited. And in terms of yield, seed cotton weight, lint weight, and lint percentage of some upland-island hybrids had MPH, but they were still significantly lower than those of upland cotton parents. [Conclusion] Two long-staple cotton hybrid combinations T035 and T044 with 5A grade high-quality were obtained, and an excellent material of G. barbadense Ta10-280 was screened. This study provides valuable data for the genetic law of fiber quality heterosis of upland-island hybrid cotton.
    Regulation of Free Amino Acid in Cotton at the Seedling Stage by Aphis gossypii Feeding
    Ma Yan, Zhang Shuai, Luo Junyu, Zhu Xiangzhen, Gao Xueke, Cui Jinjie
    2020, 32(4):  360-369.  doi:10.11963/1002-7807.mycjj.20200526
    Abstract ( 133 )   PDF (962KB) ( 27 )  
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    [Objective] Free amino acids are important nitrogen nutrients, and their changes reflect the interaction between insects and host plants. The purpose of this experiment was to study the effect of aphid feeding on the free amino acid content of host cotton in seedling stage. [Method] The seedling cotton infected by aphid with different damage grades was selected from the field and analyzed the content of free amino acid in euphylla, cotyledon, hypocotyl and root. [Result] The content of free amino acids in cotton tissues increased after feeding by Aphis gossypii, and the content of free amino acids in cotyledon, hypocotyl and root is more than 2 times of the level 0. The content of non-essential amino acids increased more significantly than that of essential amino acids. The changes of single free amino acid content in different tissues were not completely consistent, and the Asn and Glu increased significantly in all tissues. [Conclusion] The feeding behavior of cotton aphid affects the nutritional level of cotton tissue and the content of free amino acid increased significantly. 
    Study Crystallinity of the Developing Cotton Fibers by Micro-Fourier Transform Infrared Spectroscopy (FTIR) and X-Ray Diffraction (XRD)
    Zhang Lipeng, Zhang Shiding, Xu Peng, Li Xianchang, Zhang Zhen, Fan Senmiao, Gong Juwu, Yuan Youlu, Shang Haihong, Zou Huawen
    2020, 32(4):  370-380.  doi:10.11963/1002-7807.zlpzhw.20200610
    Abstract ( 118 )   PDF (5428KB) ( 23 )  
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    [Objective] The change of crystalline index (CI) of two different cultivated cotton fibers at the development stage was studied by micro-Fourier transform infrared spectroscopy (micro-FTIR) and X-ray diffraction (XRD). The feasibility was verified by measuring CI in developing cotton fibers with micro-FTIR method. And it was utilized to estimate the CI of mature cotton fibers. [Method] Upland cotton 0-153 and sea island cotton S-6 were selected as examined materials. The cotton fibers of 2 varieties were obtained at 5-30 d post anthesis respectively, with a sampling interval of 5 d. After obtaining the fiber, it was washed several times with distilled water and then placed in an oven at 40℃ for 48 hours. After drying, the FTIR and XRD spectra of each sample were obtained. The CI is calculated according to 4 different FTIR-CI calculation methods, the crystallinity changes of different cotton varieties in the developing period were compared, and the correlation between the FTIR-CI and XRD-CI was studied. [Result] Only FTIR-CI calculated by the Carrillo-Colom index(FTIR-CCI) had a good correlation with XRD-CI method, with higher R2 than 0.9 of both varieties. The fitting model (IR-CI) between the FTIR-CCI and XRD-CI was used to calculate the CI of the twenty-three randomly selected mature fibers. The results showed that the accuracy of IR-CI was good, and the XRD-CI results were within the error range of the calculated results according to the IR-CI model, while the precision of IR-CI could not reach anticipation. [Conclusion] Micro-FTIR can be used to study the change of cellulose in cotton fiber during the developing period. The model of IR-CI established by the correlation between the FTIR-CCI and XRD-CI, can be used to evaluate the crystallinity of developing period in cotton fiber. However, for the study of the crystallinity of mature fibers, it is necessary to use a large number of samples in the later experiment to establish an optimized model.