15 May 2013, Volume 25 Issue 3
    

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  • LI Xi-Hua, WU Man, YU Ji-Wen, ZHANG Jin-Fa, FAN Shu-Li, SONG Mei-Zhen, PANG Chao-You, YU Shu-Xun
    Cotton Science. 2013, 25(3): 189-196.
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    To obtain global insights into early developing fiber transcriptome characteristics, six sequencing libraries of early developing cotton fiber were constructed and sequenced using Illumina RNA sequencing. These libraries represented initiation (0 d post-anthesis (DPA) and 3 DPA) and elongation (10 DPA) stages from two backcross inbred lines having significant differences in fiber length: NMGA-062 (32.58 mm) and NMGA-105 (27.06 mm). Each sample yielded 4.6 Gb of available transcriptome data, with 98464 unigenes longer than 200 bp obtained by de novo assembly. When we compared NMGA-062 with NMGA-105 at 10 DPA, we uncovered 1931 differentially expressed genes(DEGs), of which 1536 were up-regulated and 394 were down-regulated. Gene Ontology functional enrichment and pathway enrichment analyses revealed that the DEGs were primarily associated with lipid transport and metabolism pathways, suggesting that lipid-related genes play an important role in cotton fiber elongation. The large number of DEGs detected by comparative analysis of 10-DPA cotton fiber transcriptome profiles provides a firm foundation for cloning and functional verification of fiber-related genes.
  • XING Jin, LIU Wei-Na, ZHAO Lin-Lin, ZHAO Jin-Feng, ZHANG Wen-Hui, ZHAO Bao-Hua, LI Xue-Yong
    Cotton Science. 2013, 25(3): 197-204.
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    Expression of the cotton GhTCP2 gene, a member of the TCP family, was detected by RT-PCR in young tissues, such as apical, lateral, and floral buds, of cotton plants. To study GhTCP2 gene expression patterns in detail, the GhTCP2 promoter region was isolated using the genome walking technique. Many cis-regulatory motifs related to the cell cycle, cytokinin, and auxin were identified using the on-line software program PLACE. The GhTCP2 promoter was inserted upstream of the GUS reporter gene, and the resulting plant expression construct was transformed into Arabidopsis. Histological staining of transgenic Arabidopsis plants revealed that the GhTCP2 promoter-driven GUS gene was expressed in root tips, young leaves, and apical, lateral, and floral buds. This result indicates that the cotton GhTCP2 promoter is able to drive foreign gene expression in young plant tissues. The GhTCP2 promoter has the potential to be utilized in plant insect-resistance genetic engineering to protect young plant tissue from insect attack.
  • LI Guang-Lei, FAN Shu-Li, SONG Mei-Zhen, PANG Chao-You, WEI Heng-Ling, YU Shu-Xun-
    Cotton Science. 2013, 25(3): 205-210.
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    In the present study, a WRKY gene, GhWRKY4, was cloned from cotton(Gossypium hirsutum L.). The full-length cDNA of GhWRKY4 was 1281 bp long and contained a 1083-bp open reading frame (ORF) encoding a putative group-IIc WRKY protein of 360 amino acids. Bioinformatic analysis of the GhWRKY4 gene revealed that GhWRKY4 consists of three exons and two introns. A subcellular localization assay confirmed that the protein product was targeted in onion epidermal cell nuclei. Quantitative RT-PCR indicated that GhWRKY4 is constitutively expressed in cotton roots, stems, leaves, and flowers. The 5' flanking region of GhWRKY4 was obtained by genome walking. Bioinformatic analysis revealed that GhWRKY4 contains a series of putative cis-acting elements involved in stress regulation. Further stress treatments combined with gene expression analysis indicated that GhWRKY4 is involved in regulation of salt and cold stress.
  • YANG Xin-Lei, ZHOU Xiao-Dong, LIU Heng-Wei, WANG Sheng-Fen, WU Li-Qiang, LI Zhi-Kun, ZHANG Yan, ZHANG Gui-Yin, MA Zhi-Ying
    Cotton Science. 2013, 25(3): 211-216.
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    In this study, we designed AFLP(Amplified fragment length polymorphism) markers associated with important agronomic traits using 91 BC1F2 plants derived from an interspecific cross between Gossypium hirsutum cv. CCRI 8 and G. barbadense cv. Pima 90-53. Ninety-one rows of BC1F2:3 plants were grown in the fields at Baoding suburban, Qingxian County, Hebei Province, China. The rows were scored for five yield traits-lint percentage, seed weight, boll weight, lint weight, and seed cotton weight, and five fiber quality traits-2.5% span length, micronaire, fiber uniformity, fiber elongation, and fiber strength. To provide a theoretical basis for marker-assisted selection, correlations between molecular markers and agronomic traits were tested by stepwise multiple linear regression analysis. Based on the regression analysis, 125 loci from 20 AFLP primer combinations were correlated with the 10 measured traits, although the correlations were only significant (P ≤0.05, P ≤0.01) for 15 loci from 4 primer pair combinations. In addition, six loci were associated with 6.2%-30% of the phenotypic variability observed in nine traits. Our results indicate that these AFLP markers should be useful for molecular marker-assisted selection in future cotton breeding.
  • ZHOU Zhong-Li, DU Xiong-Ming, PAN Zhao-E, HE Shou-Pu
    Cotton Science. 2013, 25(3): 217-226.
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    In this study, we used SSR markers to analyze the genetic diversity of 200 accessions of geographically varied Gossypium arboreum cultivars in the National Medium-term Gene Bank of Cotton in China. Abundant genetic diversity was uncovered. Using 83 SSR primers, we detected 368 alleles, 329 of which were polymorphic. The mean number of polymorphic alleles per locus was 3.964. PIC values ranged from 0.010 to 0.882(average = 0.578); 33 loci (39.8% of the 83 SSRs) were identified as possessing high information content(PIC > 0.7). The Shannon-Weaver diversity index(H′) ranged from 0.031 to 2.163, and the effective number of alleles(Ne) varied from 1.010 to 8.496. With respect to molecular genetic diversity, the three main cotton-growing areas of China were ordered from highest to lowest as follows: South China, the Yangtze River region, and the Yellow River region. This ranking is consistent with the widely accepted theory that G. arboreum cultivation in China spread from south to north, and the fact that South China is the most germplasm-rich region for the G. arboreum landrace 'Sinense'. A UPGMA cluster analysis was performed on the genetic distance matrix using NYSTS-pc2.20 software. Similarity coefficients among the 200 G. arboreum accessions ranged from 0.58 to 0.997(average = 0.745), with the accessions clustering into eight groups at a genetic distance of 0.73. The cultivar 'Guichi Xiaozimianbaizi' was genetically distant from the other accessions, showing specific genetic variation. No significant correlation was observed between genetic and geographic distance, although genetic distance was generally consistent with degree of geographical separation.
  • CHENG Hua, GAN Yi-Mei, LIU Fang, CAI Xiao-Yan, WANG Chun-Ying, WANG Yu-Hong, WANG Kun-Bo
    Cotton Science. 2013, 25(3): 227-233.
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    Cotton is an important commercial crop around the world, however, its chromosomes are too small and similar to identify by karyotypic analysis and chromosome banding. The cytogenetics of cotton lag behind many other crop species, and the nomenclature for individual chromosome hasn't been widely accepted except for G. hirsutum. A set of 13 BAC clones specific to A-subgenome chromosomes of G. hirsutum are labeled and hybridized with the chromosomes in metaphase of A-subgenome of G. barbadense cv. pima 90-53, G. herbaceum cv. hongxing, and G. herbaceum raced africanum by BAC-FISH. There are distinctive signals on each chromosome, so these BAC clones are proved to be cytogenetic markers to identify each of them. Most of signals are located on the terminal of chromosomes. The signals on corresponding chromosomes have similar position among the three cotton varieties. Six BAC clones are consistent with their SSR markers by comparing the physical map of BAC clones with the genetic linkage map of SSR markers, and the individual chromosome can be linked with the linkage group. The chromosomes were numbered as Ab01- Ab13 (or A(AD)■01- A(AD)■13) in A-subgenome of G. barbadense, A101- A113 in G. herbaceum, and A1a01- A1a13 in G. herbaceum raced africanum. The results will help to analyze the origin and evolution of cotton, to orient new genetic markers on chromosomes, to construct physical map of chromosomes, and so on.
  • WANG Fei, KUANG Meng, XU Hong-Xia, WANG Yan-Qin, ZHOU Da-Yun, FENG Xin-Ai, FANG Dan, XING Yao-Fei, YANG Wei-Hua-
    Cotton Science. 2013, 25(3): 234-239.
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    In an SSR study of seven conventional cotton cultivars, we analyzed 24 seeds of each cultivar using 52 core primer pairs. Among the seven cultivars, the number of miscellaneous seeds ranged from 1 to 9, and averaged 3.9. The percentage of homozygous DNA loci varied from 92.3% to 100%, and averaged 97.0%. Our analysis revealed that varietal purity and DNA locus homozygosity levels were highly positively correlated. Using SSR molecular markers, we were able to accurately differentiate between the impacts of genetic factors and seed production procedures on cultivar purity.
  • PAN Li-Dong, SHI Ming, ZHANG Kai, CHEN Jin, GAO Cong-Fen
    Cotton Science. 2013, 25(3): 240-246.
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    F1 screening method is the best bioassay measure to detect the resistance allele frequency of Helicoverpa armigera (Hübner) to transgenic Bt cotton if there is a high Bt-resistant strain. From 2010 to 2012, the frequency of allele conferring resistance to Bt cotton in Qiuxian County, Hebei Province, was estimated by using the F1 screening method. The results showed that from 2010 to 2012, 32 out of 122, 22 out of 141 and 22 out of 141 field-collected male moths were detected to carry resistance alleles and the resistance allele frequency was estimated as 0.131 (95% confidence interval (CI): 0.101-0.162), 0.078 (95% CI: 0.034-0.122) and 0.199 (95% CI: 0.124-0.274), respectively. Bioassay of Helicoverpa armigera populations on Bt diet was adopted to detect resistance to Cry1Ac in 2012. The results showed that the resistance of field population from Qiuxian County was the highest, the resistance ratio reached 19.2 fold, compared with susceptible strain, and the resistance ratio of other three populations which ranged between 4.9 and 9.3 fold. Long-term adoption of planting uni-toxin-producing Bt cotton, and lack of conventional cotton refuge system may accelerate the resistance evolution in this region, and it is necessary to establish and implement effective resistance management strategy as soon as possible.
  • ZHANG Jian-Hong, GUO Bao-Sheng, ZHAO Gui-Yuan, GENG Jun-Yi, CUI Rui-Min, WANG Zhao-Xiao, LIU Cun-Jing, LIU Su-En, ZHANG Xiang-Yun
    Cotton Science. 2013, 25(3): 247-253.
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    We analyzed genetic diversity of 48 cotton varieties using 48 SSR (Simple sequence repeat) primer pairs for fiber quality QTLs(Quantitative trait loci) broadly distributed across chromosomes 7, 13, and 25. Of the 48 primer pairs, 3 yielded polymorphic bands. Correlation analysis indicated that the corresponding loci were significantly associated with fiber length and strength. Comparison with characteristic bands of Gossypium barbadense (Sea island cotton) indicated that some useful cotton quality alleles were derived from that species. These results may serve as a resource for molecular marker-assisted selection pyramiding of favorable traits in future breeding programs.
  • CHEN Bing, WANG Fang-Yong, HAN Huan-Yong, LIU Zheng, DENG Fu-Jun, LIN Hai, YU Yu, LI Shao-Kun, WANG Ke-Ru, XIAO Chun-Hua
    Cotton Science. 2013, 25(3): 254-261.
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    Using data from red-edge parameters of haperspectra, we endeavored to provide an expedient way to extract leaf nitrogen contents(LNC) of cotton infected with Verticillium wilt, and to lay the groundwork for estimating cotton yield infected by Verticillium wilt using remote sensing technology. The relationship between LNC and red edge parameters were analyzed, and diagnose models of spectra red edge parameters were established for cotton leaves infected by Verticillium wilt. The main results are as follows: (1) With the increase of leaf severity level, LNC little by little decreased, difference is significant. (2) In all red edge parameters, REP, Dr, Lo, Depth672 and Area672 all decreased, and the degree of decrease was maximum to Area672 value, the degree of decrease was minimum to Dr value; but  Lwidth largely increased. (3) LNC had best significant positive correlations with REP, Lo, Depth672 and Area672, had best significant negative correlations with Lwidth, and no best significant correlations with Dr. (4) Diagnose models of LNC to disease cotton leaves on the basis of spectra red edge parameters all attached best significant correlations (P<0.01). The diagnose models based on Area672 had best estimated precision for LNC, and R2 were over 0.7, RMSE less than 0.6, RE  less than 0.007. So using red edge parameters of haperspectra monitor LNC of cotton infected by   Verticillium wilt is accurate and convenient, and those models can better diagnose LNC.
  • LI Cai-Hong, LI Zhi-Fang, FENG Zi-Li, ZHAO Li-Hong, SHI Yong-Qiang, WANG Ling-Fei, LIU Yi-Jie, ZHU He-Qin
    Cotton Science. 2013, 25(3): 262-268.
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    With many fungal genomes having been or currently being sequenced, identification of gene function has become a hot topic in fungal research. Gene knockout is one of several techniques used in studies of gene function. The method can be used to delete entire genes, replace an allele with another, exchange a given gene's normal promoter for a regulatable one, and label genes with epitope tags or fluorescent proteins. In this paper, we review applications of gene knockout via Agrobacterium tumefaciens-mediated transformation (ATMT) in filamentous fungus functional genomics.
  • WU Xia, LI Yan-E, SHANG Guan-Xiao-Xia
    Cotton Science. 2013, 25(3): 269-277.
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    Transcription factors play important regulatory roles in various stages of cotton fiber development, beginning with initiation and continuing through secondary cell wall synthesis and maturity. In recent years, many transcription factors related to cotton fiber development have been reported, including MYB, HD-ZIP, MADS, and TCP families. Among them, MYB transcription factors have been identified as essential proteins involved in this process. Arabidopsis trichome development, which has been well characterized at the molecular level, is regulated by a MYB-bHLH-WD40 protein complex. As several studies using cotton fiber-related genes have demonstrated a close relationship between cotton seed fibers and Arabidopsis leaf trichomes, cotton fiber development and Arabidopsis leaf trichome differentiation likely share similar regulatory mechanisms. MIXTA-like MYB genes such as GhMYB25 and GhMYB25-like also have significant regulatory roles in cotton fiber development. GL1-like MYB genes and MIXTA-like MYB genes most likely control cotton fiber development through different regulatory networks. Extensive research on transcription factors related to cotton fiber cells has provided insights into the molecular mechanisms of fiber cell differentiation and development. This paper discusses some achievements in this field.
  • LI Peng-Cheng, DONG He-Lin, LIU Ai-Zhong, LI Ru-Yi
    Cotton Science. 2013, 25(3): 278-282.
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    Field experiment was conducted to study the effect of exogenous hormones on cotton(Gossypium hirsutum L. cv SCRC 28) growth and development during the early growth period. Six different concentrations of three exogenous hormones were designed, which were heteroauxin(IAA) 5 mg·L-1 and 10 mg·L-1, 6-benzyl adenine (6-BA) 50 mg·L-1 and 100 mg·L-1, gibberellin acid(GA) 20 mg·L-1 and 50 mg·L-1, with water as control, for foliar application three times from seedling stage(6-7 true leaves) to squaring stage. The results showed that the vertical growth rates of main stem of cotton plants with six different concentrations of exogenous hormones by foliar application during seedling stage were all slower than those of control. Diameters at cotyledonary node of main stem of cotton plants with six exogenous hormones treatments were greater than those of control. The growth rates of leaves of cotton plants were significantly speeded up by treatment of 50 mg·L-1 GA, with the leaf areas of whole plants increasing significantly. SPAD values of chlorophyll in functional leaves(4th leaf from the top) of cotton plants with six different concentrations of exogenous hormone treatments were significantly higher than those of control. Treatments with different concentrations of IAA, GA promoted the development of bud of cotton plants, and the treatment of 20 mg·L-1 GA achieved marked effect. The results above showed that in the experimental conditions vegetative and reproductive growth of cotton plant could be coordinated and buds could be better developed from seedling stage to squaring stage by foliar spraying IAA with concentrations of 5 mg·L-1,10 mg·L-1 or GA with concentrations of  20 mg·L-1, 50 mg·L-1. Effect of GA was better than that of IAA, and effect of 20 mg·L-1 GA was the best among 4 treatments. Treatments with 6-BA was noneffective to promote the development of buds.