Characterization of Gh SERK2 and its expression associated with somatic embryogenesis and hormones level in Upland cotton

Somatic embryogenesis （SE） is one of the most important steps during regeneration of cotton, but the molecular mechanism of SE remains unclear. SOMATIC EMBRYOGENSIS RECEPTOR KINASE （SERK） gene is known to function in SE. A homolog GhSERK2 （accession number： JF430801） was cloned from Upland cotton and characterized for its functions in SE. GhSERK2 expressed in different tissues and showed higher expression level in floral organs than vegetative ones with the highest levels in ovule and anther. GhSERK2 expressed during SE with a high level at globular embryos stage. Upon treatment with indole-3-butytic acid （IBA）, the transcription level of GhSERK2 was induced and promoted SE subsequently. A 2-day treatment of 2,4-dichlorophenoxyacetic acid （2,4-D） induced the expression of GhSERK2, but treatments of 2,4-D for longer periods sharply inhibited the GhSERK2 transcription level of embryogenic callus （EC）. The levels of hormones, including 3-indoleacetic acid （IAA）, abscisic acid （ABA）, and brassinosteroid （BR）, were increased in the initial calli induced from the over-expression of GhSERK2 cotton. Our results indicated that GhSERK2 expression was associated with induction of SE and closely related to hormone levels during tissue culture in Upland cotton, and the gene might play an important role in regeneration of cotton.

Intron-Targeted Intron-Exon Splice Conjunction (IT-ISJ) Marker and Its Application in Construction of Upland Cotton Linkage Map

To develop a new DNA maker, which could be used in genetic diversity analysis and genetic map construction in plants, IT-ISJ （intron targeted intron-exon splice junction） primer combinations, which were designed according to the intronexon splice junction conserved sequences, were used to construct cotton genetic linkage map in the present study. 49 out of 704 IT-ISJ primer combinations showed polymorphism between upland cotton high quality cultivar Yumian 1 and multiple dominant gene line T586, and the polymorphic primer combinations accounted for 7.0% of total primer combinations. 49 IT-ISJ primer combinations were used to genotype 270 F2：7 recombinant inbred lines developed from （Yumian 1 × T586） F2, and 58 IT-ISJ loci were obtained. 58 IT-ISJ, together with 150 SSR and 8 morphological loci, were used to conduct linkage analysis, and a linkage map including 22 linkage groups and 113 loci （49 IT-ISJ, 62 SSR, and 2 morphological loci） was constructed. The linkage map covered 714.5 cM with an average interval of 6.3 cM between two markers, accounting for 16.1% of cotton genome. The present study demonstrated that the polymorphism of IT-ISJ marker is high, and it could be effectively applied in plant genetic map construction.

Selection and Characterization of a Novel Glyphosate Tolerant Upland Cotton(Gossypium hirsutum L.) Mutant(R1098)

Stepwise selection approach was adopted to obtain glyphosate-tolerant upland cotton mutant(R1098) from the embryogenic calli of Coker 312(Gossypium hirsutum L.).The calli were transferred to selection medium and multi-step selection pressure process was carried out until the

An Integrated Genetic,Physical and Transcript Map of Homoeologous Chromosomes 12 and 26 in Upland Cotton(Gossypium hirsutum L.)

While Upland cotton(Gossypium hirsutum L.) represents 95% of the world production,its genetic improvement is hindered by the shortage of effective genomic tools and resources.The

Biochemical Genetic Mechanism and QTLs of Early Maturing without Yield Loss in Short-season Upland Cotton(Gossypium hirsutum L.)

The short season cotton(SSC) was important Upland plant ecotype(Gossypium hirsutum L.).The growth of SSC was very short that is 105 ～ 110 days(after planting). SSC could increase

Genome-wide association study of micronaire using a natural population of representative upland cotton(Gossypium hirsutum L.)

Background: Micronaire is a comprehensive index reflecting the fineness and maturity of cotton fiber.Micronaire is one of the important internal quality indicators of the cotton fiber and is closely related to the value of the cotton fiber.Understanding the genetic basis of micronaire is required for the genetic improvement of the trait.However,the genetic architecture of micronaire at the genomic level is unclear.The present genome-wide association study(GWAS)aimed to identify the genetic mechanism of the micronaire trait in 83 representa:tive upland cotton lines grown in multiple environments.Results GWAS of micronaire used 83 upland cotton accessions assayed by a Cotton 63 K Illumina Infinium single nucleotide polymorphism(SNP)array.A total of 11 quantitative trait loci(QTLs)for micronaire were detected on 10 chromosomes.These 11 QTLs included 27 identified genes with specific expression patterns.A novel QTL,qFM-A12–1,included 12 significant SNPs,and GhFLA9 was identified as a candidate gene based on haplotype block analysis and on strong and direct linkage disequilibrium between the significantly related SNPs and gene.GhFLA9 was expressed at a high level during secondary wall thickening at 20∼25 days post-anthesis.The expression level of GhFLA9 was significantly higher in the low micronaire line(Msco-12)than that in the high micronaire line(Chuangyou-9).Conclusions: This study provides a genetic reference for genetic improvement of cotton fiber micronaire and a foundation for verification of the functions of GhFLA9.

Development of glyphosate-tolerant transgenic cotton plants harboring the G2-aroA gene

Given that glyphosate weed control is an effective strategy to reduce costs and improve economic outcomes of agricultural production in China,the development of glyphosate-resistant cotton holds great promise.Using an Agrobacterium-mediated transformation method,a new G2-aroA gene that encodes 5-enolpyruvylshikimate-3-phosphate synthase（EPSPS）was transformed into cotton cultivar K312.The transgenic cotton plants were regenerated from a callus tissue culture via kanamycin selection.Ten regenerated cotton plants were obtained and allowed to flower normally to produce fruit.The results from polymerase chain reaction（PCR）and Southern and Western blot analyses indicated that the target gene was integrated into the cotton chromosome and was expressed effectively at the protein level.The glyphosate tolerance analysis showed that the transgenic cotton had a high resistance to glyphosate.Further,even cotton treated with 45.0 mmol L^–1 of glyphosate was able to slowly grow,bloom and seed.The transgenic cotton may be used for cotton breeding research of glyphosate-tolerant cotton.

Comparative Analysis of Salinity-Induced Proteomic Changes in Cotton (Gossypium hirsutum L.)

Salt stress on cotton varieties of distinct salinity tolerance can induce expression of different proteins. Zhong 07, a salt-tolerant variety and Zhong s9612, a salt-sensitive variety, were utilized as experimental materials. The leaves of trefoil seedlings treated with or without 0.4% NaCl for 24 h were harvested for whole-protein extraction. Two-dimensional technology, combined with mass spectroscopy (MS) analysis and protein database searching, was employed to detect differentially expressed proteins and determine their identities and biological functions. Compared with the control, Zhong 07 showed 10 differentially expressed proteins under salt stress, of which 6 were upregulated and 4 were downregulated. Meanwhile, 12 differentially expressed proteins were detected in Zhong s9612 under salt stress, of which 10 were upregulated and 2 were downregulated. In the matrix-assisted laser desorption-ionization/time of flight-time of flight/MS analysis, 14 differentially expressed proteins were successfully identified, including the ribulose-1, 5-bisphosphate carboxylase/oxygenase (RuBisco) large subunit-binding protein subunit alpha (RuBisco α), luminal binding protein (LBP), heat shock protein 70 (Hsp1, 2, 3), pathogenesis-related protein class 10 (PR-10), quinoneoxidoreductase-like protein (QOR), S-adenosylmethioninesyn-thetase (SAMS), enolase (EN), and RuBisco large subunit-binding protein subunit beta (RuBisco β). Cellular function is ultimately executed by proteins, and cotton varieties with different salt tolerance can be influenced by salt stress to various degrees, which can provide certain theoretical foundation for the identification of salt tolerance of cotton varieties. The findings also provide some proteins, such as the RuBisco large subunit binding proteins α and β subunits, OEE2 protein, HSP70, and S-adenosylmethionine synthetase, which can be used as protein markers of salt-to-lerance before- and post-treatment, making a big difference in salt-tolerance identification in cotton.

Comparative Transcriptome Analysis of Seed Germination of a Cotton Variety with High Tolerance to Low Temperature

Gossypium hirsutum L.is an important cash crop native to the subtropics and is widely cultivated around the world.Low temperature is an important stress that seriously affects seed germination and emergence during planting.In this study,transcriptomic profiles of low-temperature-and normal-temperature-germinated seeds of Xinluzao 25,a variety with low-temperature tolerance and high germination rates,were analyzed and compared.The following results were obtained.(1)A total of 81.06 Gb of clean data were obtained after transcriptome sequencing and assembly,and 76,931 non-redundant Unigene sequences were obtained after data consolidation and concatenation;of these,69,883 Unigene sequences were annotated.In addition,55,463 Unigene transcript sequences(72.2%)were annotated for Gene Ontology(GO)classification,and 26,629 genes were involved in 50 metabolic pathways identified by Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis.(2)Three main pathways related to low-temperature tolerance of seed germination were identified:starch and sucrose metabolism,phenylpropanoid biosynthesis,and cysteine and methionine metabolism.Their main molecular functions involve the regulation of abscisic acid and activities of enzymes such as amylase,peroxidase,and oxidoreductase.During germination at low temperature,more genes were down-regulated than up-regulated genes at the protrusion stage(2 mm),and more genes were up-regulated than down-regulated at the germination stage(30 mm)after protrusion.(3)The enzyme activities at the two stages showed that amylase,peroxidase,catalase,and glutathione reductase had higher activities when the seeds germinated at 15℃.In this study,high expression of amylase,peroxidase,catalase,and glutathione reductase genes may be the main cause of increased tolerance to low temperature.

多环境下陆地棉(Gossypium hirsutum L.)重组自交系铃重与衣分性状的QTL分析

本研究利用以SGK9708为母本,0-153为父本构建的196个陆地棉重组自交系(F6:8)构建了包含186个标记,总长827.84cM,标记间平均距离4.45cM,覆盖棉花基因组18.6%的遗传连锁图谱,并对7个环境下的铃重和衣分性状进行QTL定位和上位性互作分析。利用两种分析软件(WinQTLcart2.5和QTLNetwork2.0)共同定位了多个环境下稳定表达的5个主效QTLs(qBW-1-1,qBW-1-2,qLP-2-1,qLP-2-2和qLP-4-2)。利用QTLNetwork2.0分别检测到4对铃重上位性互作QTLs和7对衣分上位性互作QTLs,以背景位点间的互作和加性效应位点与背景位点间的互作为主。除主效QTL外,上位性效应也是陆地棉铃重和衣分性状的重要遗传基础。本研究定位的5个主效QTLs为选择高铃重、高衣分品种的棉花分子标记辅助育种提供了重要依据。

陆地棉吐絮铃数及吐絮率的QTL定位

采用复合区间作图法，对陆地棉sGK9708X0-153组合的重组近交系（RI）及永久F2群体在曲周（2009年）、安阳（2009，2010年）3个环境条件下的吐絮铃数和吐絮率进行QTL检测，共获得18个QTLs。其中，7个与吐絮铃数相关，加性效应在-0．46～0．33之间，可解释的表型变异为5．86％～11．28％；11个与吐絮铃率相关，加性效应在-3．64％～3．20％之间，可解释的表型变异4．68％-9．84％。这些QTLs主要分布在25号（吐絮铃数／吐絮率：3个／6个，下同）、16号（2个／2个）、18号（1个／2个）染色体和LG49（1个／1个）上。这18个QTLs中，qPOB-16-2在R1群体中的3个环境下被稳定检测到，qCOB．16．1、qPOB-25—3和qPOB-25．4在RI中的两个环境下被稳定检测到。这些QTLs可以应用于吐絮铃数和吐絮率的分子标记辅助选择。

QTL mapping for fiber quality and yieldrelated traits across multiple generations in segregating population of CCRI 70

Background:Cotton is a significant economic crop that plays an indispensable role in many domains.Gossypium hirsutum L.is the most important fiber crop worldwide and contributes to more than 95%of global cotto n production.Identifying stable quantitative trait locus(QTLs)controlling fiber quality and yield related traits are necessary prerequisites for marker-assisted selection(MAS).Results:A genetic linkage map was constructed with 312 simple sequence repeat(SSR)loci and 35 linkage groups using JoinMap 4.0;the map spanned 1 929.9 cM,with an average interval between two markers of 6.19 cM,and covered approximately 43.37%of the cotton genome.A total of 74 QTLs controlling fiber quality and 41 QTLs controlling yield-related traits were identified in 4 segregating generations.These QTLs were distributed across 20 chromosomes and collectively explained 1.01%?27.80%of the observed phenotypic variations.In particular,35 stable QTLs could be identified in multiple generations,25 common QTLs were con sistent with those in previous studies,and 15 QTL clusters were found in 11 chromosome segments.Conclusion:These studies provide a theoretical basis for improving cotton yield and fiber quality for molecular marker-assisted selection.

Cloning and Expression Analysis of Chalcone Synthase and Chalcone Isomerase Encoding Genes in Gossypium hirsutum

In this study,chalcone synthase and chalcone isomerase encoding genes(GhCHS and GhCHI)were cloned from upland cotton(Gossypium hirsutum)cultivar HM-40,and analyzed bioinformatically.Their functions were analyzed through virus induced gene silencing(VIGS).The results showed that GhCHS gene has an open reading frame(ORF)of 1170 bp and encodes a protein of 389 amino acids.Many phosphorylation sites were detected in GhCHS protein,suggesting that it may be involved in kinase phosphorylation.The deduced GhCHS protein was most closely related to Theobroma cacao CHS protein according to phylogenetic analysis.The GhCHI ORF was 630 bp and encoded a protein of 209 amino acids.Many phosphorylation sites were found in GhCHI protein,indicating that it may be related to kinase phosphorylation.The GhCHI protein was most closely related to Hibiscus cannabinus CHI protein according to phylogenetic analysis.Quantitative PCR(q PCR)showed that GhCHS and GhCHI were rapidly activated after inoculation with Verticillium dahliae VD07,and then their expression levels kept increasing over time,indicating that the two genes might play an important role in the defense response against Verticillium wilt.Virus induced gene silencing(VIGS)was used to silence endogenous GhCHS and GhCHI genes in upland cotton plants before VD07 was inoculated to identify disease resistance.The results showed that the disease index of plants untreated with Agrobacterium tumefaciens was 31.2,and that of the plants inoculated with empty vector was 30.0.The disease index of GhCHS-silenced plants was 72.5,and that in GhCHI-silenced plants was 67.5.These results confirmed that GhCHS and GhCHI may play an important role in defense response of upland cotton to Verticillium wilt.

Study on the Stability of World Diversity of Cultured Species <i>G. hirsutum</i>L. to Salination

The paper presents the results of a study of salt tolerance in some different eco-geographical samples of the cotton germplasm collection of the Institute of Genetics and Experimental Biology of the Academy of Sciences of the Republic of Uzbekistan. According to the results obtained, the studied samples were divided into several groups depending on their salt tolerance. Salt tolerant and unstable samples were found in all studied ecological and geographical groups, but differed in the frequency of distribution.

Construction of a linkage map and QTL mapping for fiber quality traits in upland cotton (Gossypium hirsutum L.)

With the development in spinning technology, the improvement of cotton fiber quality is becoming more and more important. The main objective of this research was to construct a high-density genetic linkage map to facilitate marker assisted selection for fiber quality traits in upland cotton (Gossypium hirsutum L.). A genetic linkage map comprising 421 loci and covering 3814.3 cM, accounting for approximately 73.35% of the cotton genome, was constructed using an F2 population derived from cross GX1135 (P 1 )×GX100-2 (P 2 ). Forty-four of 49 linkage groups were assigned to the 26 chromosomes. Fiber quality traits were investigated in F2 population sampled from individuals, and in F2:3 , and F2:4 generations sampled by lines from two sites and one respectively, and each followed a randomized complete block design with two replications. Thirty-nine quantitative trait loci were detected for five fiber quality traits with data from single environments (separate analysis each): 12 for fiber length, five for fiber uniformity, nine for fiber strength, seven for fiber elongation, and six for fiber micronaire, whereas 15 QTLs were found in combined analysis (data from means of different environments in F2:3 generation). Among these QTLs, qFL-chr5-2 and qFL-chr14-2 for fiber length were detected simultaneously in three generations (four environments) and verified further by combined analysis, and these QTLs should be useful for marker assisted selection to improve fiber quality in upland cotton.

陆地棉超矮杆突变体基因的初步定位

陆地棉超矮杆是一种新发现的突变体材料。前期对其遗传规律的研究表明,超矮杆突变性状是受一对完全隐性基因du控制的质量性状。文章以"超矮1号"和"新陆早16"配置的F2为作图群体,通过对双亲以及近等基因池的筛选,在1 350对SSR引物中共获得70对多态性引物。而后检测F2作图群体每个单株的基因型,显示有36个标记可以连锁,分布在8个连锁群上,其中超矮杆突变性状位于连锁群LG01。与目标基因du连锁的分子标记有7个:NAU2679、NAU2749、NAU905、NAU2838、NAU5373、NAU2238和NAU4946,它们皆为共显性标记。依据现有的棉花遗传图谱,标记NAU4946、NAU2238、NAU905、NAU5373和NAU2679位于第6染色体上,而目标基因du位于NAU2238和NAU4946之间,遗传距离分别为3.3 cM和1.4 cM,由此推定du在第6条染色体上。

陆地棉纤维品质和产量性状的遗传分析

两个陆地棉型纤维品质优异品系９４１７３和９４２５０分别与推广品种中棉所１２号、鲁棉１１号、中棉所１６号和鲁棉１０号杂交，按方法２模式Ⅰ配成６×６共２１个组合。纤维长度、马克隆值、籽棉产量、单铃重和籽指的Ｇ．Ｃ．Ａ．效应达显著水平，除衣分率外，上述各性状Ｓ．Ｃ．Ａ．效应均达显著水平。其中纤维长度、马克隆值和籽指Ｇ．Ｃ．Ａ．方差／Ｓ．Ｃ．Ａ．方差比值较大，证明以加性效应为主。Ｇ．Ｃ．Ａ．估算结果表明，９４１７３和９４２５０是提高陆地棉品种纤维品质性状育种非常有价值的亲本，通过杂交可同时兼顾后代纤维长度、强度和细度的改良。通过Ｓ．Ｃ．Ａ．的估算认为，在某些组合Ｆ１会有杂种优势出现，而在系谱选择中应注意其干扰。在纤维长度、马克隆值、籽棉产量、单铃重、衣分率和籽指的遗传模式推导中，只有纤维长度和衣分率符合加性＋显性模式。纤维长度为部分显性，长绒基因既有隐性，也有显性，广义遗传力为７８１７％，狭义遗传力为６７４４％；衣分率广义遗传力和狭义遗传力分别只有２６０８％和２１１６％，高衣分率基因较多的表现为隐性。

陆地棉种子性状直接效应和母体效应的遗传分析

采用二倍体种子遗传模型及其分析方法,分析了陆地棉（Gossypium hirsutum L.) 5个品种及其配制的F_1、F_2和正反回交一代（BC_1;、BC_2、RBC_1和RBC_2）世代籽指、种仁率、容重和油分4个种子性状的直接效应、母体效应和细胞质效应。分析结果表明,4个棉子性状的遗传均以母体效应为主,种子直接遗传效应作用较小。各性状的直接效应与母体效应的协方差不显著。通过母体植株的遗传表现对这些性状选择更为有效。可以根据籽指小和容重大等性状母体植株的遗传表现间接选择高油分材料。母体加性效应大的亲本,其杂种后代的母体加性效应总值一般也较大。

棉花优异纤维品质性状的双列杂交分析(英文)

利用 5个具有不同纤维品质性状的品种 (系 )配制完全双列杂交组合 2 0个 ,通过亲本和F1的 2年随机区组试验 ,结果为除纤维整齐度受环境等因素影响较大 ,其余性状主要受遗传因素控制 ;在与环境的互作中 ,纤维强度和长度的互作效应小 ,麦克隆值的加性和母体效应及伸长率的显性效应与环境的互作较大 ,均达到了显著水平遗传主效中 ,所有的研究性状不存在母体效应 ,以加性为主 ;强度与长度加性遗传率高 ,分别占 77 6 %和 73 2 % ;麦克隆值的加性效应占 4 5 2 % ,显性效应所占的比例在纤维性状中最高 ,为 11 5 %。纤维品质性状的群体平均优势仅麦克隆值的较高 (3 2 % ) ,达到了显著水平 ,其余性状的优势仅为 - 0 4 %～ 0 7%。纤维品质性状的遗传结果与杂种优势一致。在杂种优势利用时 ,可以通过双亲平均值的高低来预测F1的纤维品质表现。纤维强度、长度和细度的加性遗传率高 。

陆地棉花粉及柱头生活力的研究

以11个陆地棉品种(系)为亲本,组配成7个组合,在8:30至17:30的10个时间段,分别给母本柱头授以自然花粉及冰箱保存花粉,以杂交结实特性为考查指标,研究棉花花粉及柱头生活力表现。分析认为,当地自然条件下,柱头活力为1.5d,开花当天柱头活力呈"不对称双峰"特征,9:30～10:30及15:30～16:30是柱头活力旺盛时段。花粉活力可维持1d,以9:30～10:30生活力较强,随时间推后,花粉活力大幅度衰减。依据本试验结果,就棉花杂优利用制种工作的授粉方法及技术进行讨论。