Genetic analysis of earliness in upland cotton ( Gossypium hirsutum L.). II. Yield and lint percentage

Inheritance and interrelationships of seed cotton and lint yields were evaluated in a diallel analysis involving seven early maturing parents of different origin and a commercial variety. Lint yield showed relatively little additive variance and low heritability, whereas lint percentage showed the opposite. Highest yields were shown by the least determinate and slowest-maturing genotypes; yields generally decreased as determinacy increased and rate of maturity accelerated. Except for date for first open boll, components of earliness showed no associated with yield.     

Cytogenetic analysis of translocations in cotton

In order to create cotton translocation lines, 191 translocations were recovered as heterozygotes following combined treatment of seeds by colchicine and γ‐rays, irradiation of seeds by thermal neutrons and c‐irradiation of pollen. Cytogenetic analysis showed that chromosome translocations involving two chromosomes arose more often (180) than those involving three chromosomes (11). The heterozygous translocations were characterized by different formation and frequencies of the multivalents at metaphase I of meiosis. Only 104 translocations were characterized by a multivalent frequency of more than 0.25 per cell. Most of the translocations (156) exhibited a high meiotic index, but 11 were characterized by a decreasing meiotic index and by an increase in the percentage of tetrads with micronuclei. The pollen fertility of translocations differed significantly and varied from high fertility to pollen sterility. The high frequency of abortive pollen grains was typical for about one half of translocations studied. The translocations were made homozygous and as a result, 12 new reciprocal homozygous translocation lines were obtained in cotton.     

Inheritance of time of flowering in upland cotton under natural conditions

Time to flowering is an essential component of the adaptation and productivity of cotton ( Gossipium hirsutum ) in various agro-ecological zones. This article presents a study of the genetic control of this trait in two crosses obtained from different early-maturity parental lines. In each cross, multiple generations including P₁, F₁, P₂, B₁, B₂ and F₂ were evaluated under two natural field conditions in 2004 and 2005. The data on time to flowering in the F₂ populations had a continuous distribution but deviated from normality. A joint segregation analysis (JSA) revealed that time of flowering in upland cotton was controlled by a mixture of an additive major gene and additive-dominant polygenes. The first- and second-order genetic parameters were all calculated based on the mixture of major gene and polygene inheritance models using JSA. These results suggested that there was considerable genetic diversity and complexity in days to anthesis in upland cotton. This variation can be used to formulate the most efficient breeding strategy and to design cotton for a particular environment.     

Mapping of quantitative trait loci controlling cotton leaf curl disease resistance in upland cotton

Cotton leaf curl disease (CLCuD) is a major threat to cotton production in Asia and Africa. Using marker-assisted breeding can be the best sustainable approach to tackle CLCuD. Identification of new QTLs in the indigenous cotton germplasm is necessary to combat CLCuD. The current study was designed to construct a genetic linkage map of bi-parental F2:F3 populations developed from highly tolerant MNH-886 and highly susceptible S-12 cotton cultivars. One hundred seven CLCuD-associated simple sequence repeat (SSR) marker alleles were identified as polymorphic and three new QTLs were found on chromosomes C11, C19 and C21. Two QTLs on chromosomes C11 and C19 were detected in both F2 and F3 populations in the region flanked by SSR markers CIR316 and BNL4094, and BNL285 and BNL3348, respectively. Whereas, one QTL on chromosome C21 was detected in the region flanked by SSR markers JESPR158 and JESPR135 in both F2 and F3 generations. The CLCuD-associated QTLs identified in this study can help fine-tune the molecular mapping of the QTLs on the cotton genome against CLCuD.     

Diallel analysis of root-knot nematode resistance based on galling index in upland cotton

The southern root‐knot nematode (RKN) [Meloidogyne incognita (Kofoid and White) Chitwood) RKN] is one of the most destructive pests in the Cotton Belt of the U.S. Developing and employing resistant cultivars is the most economical and efficient method for RKN management. This greenhouse investigation was conducted to understand the quantitative genetic basis of RKN resistance in the major RKN resistance sources using a nine‐parent diallel analysis based on improved RKN evaluation techniques. The selected genotypes consisted of three ‘Aubrun 623 RNR’ (here RNR = R oot‐knot N ematode R esistance)‐derived resistant lines, one moderately resistant cultivar ‘Acala Nem‐X’, and five susceptible cultivars. Comparison between F₁ and their parents in galling index revealed that the RKN resistance is partially dominant. The general combining ability was more important than the specific combining ability for RKN resistance. The estimates for broad‐sense and narrow‐sense heritabilities on galling index were 0.82 and 0.65, respectively, indicating that RKN resistance in the Auburn and Nem‐X resistance sources is largely controlled by genetic and additive effects. The minimum number of genes for RKN resistance was estimated to be two. Therefore, under reliable and uniform RKN inoculation and infestation conditions, single plant selection should be efficient in transferring Auburn RKN resistance into elite genetic backgrounds.     

新型陆地棉细胞质雄性不育系花器形态学和细胞学观察

 采用石蜡切片法,对一种来源于陆地棉的新型细胞质雄性不育材料的3份转育材料（G22A,H109A和1793A）的花药发育过程进行了花器形态及细胞学观察。花器形态和细胞学观察结果表明,G22A,H109A和1793A三个不育系均花瓣小,花丝短,柱头明显外露,败育彻底,无花粉。败育时期主要发生在造孢细胞增殖时期至小孢子母细胞减数分裂前,表明这3份不育材料不育性状不受细胞核基因的影响。对不育材料与其保持系细胞学观察发现,不育材料与正常花药发育细胞学形态上主要差别是绒毡层发育。不育系绒毡层的着色不明显,大小与中层细胞相似且在整个花药发育过程中不降解,而保持系绒毡层的着色深,在小孢子发育过程中逐渐降解。因此,根据细胞学研究结果可以说明花药中绒毡层发育异常是该来源于陆地棉的新型细胞质雄性不育材料败育的细胞学基础。     

棉花芽黄突变体10个叶绿体蛋白编码基因RNA编辑位点的测定及分析

 RNA编辑是陆生植物叶绿体转录后基因表达调控的一种方式。已有研究表明植物黄化或白化可能与叶绿体RNA编辑有关。本实验采用PCR,RT-PCR及测序的方法,确定棉花芽黄突变体V1子叶与真叶的10个叶绿体蛋白编码基因中各有34个编辑位点,有6个位点存在编辑效率的差异。将这些编辑位点与柯字310比较发现,芽黄突变体多5个编辑位点。运用生物信息学软件对34个编辑位点进行分析,结果表明accD-109, clpP-187, ndhB-50, ndhB-196, ndhD-128, ndhD-225等13个位点会影响相应蛋白二级或三级结构,表明上述编辑位点的改变可能对其蛋白的正确组装及功能的发挥起重要作用。     

Inheritance of fiber quality and lint yield in a chemically mutated population of cotton

The narrow germplasm base of the upland cotton (Gossypium hirsutum L.), grown on the Texas high plains historically, has limited improvement of fiber quality. Chemical mutagenesis and subsequent selection have helped the development of lines with improved fiber quality in cultivars adapted to this region. This study was conducted to determine the inheritance of improvements in fiber quality. M₃ lines with divergent fiber properties of micronaire, length, and strength were selected from a population of Paymaster HS 200 treated with 3% v/v ethyl methanesulfonate (EMS) for two hours. The 115 selected lines of M₄ and M₅ generation were evaluated for fiber quality and lint yield. Regression of the M₄ and M₅ on the M₃ generation, as well as the M₅ on the M₄ was used to generate narrow sense heritability coefficients. Significant variations were observed between the mutant lines in all generations except for lint yield in the M₅ (1997). The highest heritability estimates were found in fiber length (h ²= 0.29** to 0.46**). Micronaire and strength showed intermediate heritability estimates of h ²= 0.14 to 0.19, while lint yield had a very low heritability estimate of h ²= 0.03. Fiber length and strength were correlated (r= 0.58** to 0.46**) in all the three generations. The mutants identified in these studies have the potential to improve fiber quality of upland cotton without introducing alien genes that may reduce adaptation to short growing season production regions.     

棉花纤维品质不同世代的遗传相关分析

400716 重庆市西南大学农学与生命科学学院     

Inheritance of resistance to Verticillium wilt (Verticillium dahliae) in cotton (Gossypium hirsutum L.)

Verticillium wilt, caused by Verticillium dahliae Kleb., is a major constraint to cotton production in almost all countries where cotton is cultivated. Developing new cotton cultivars resistant to Verticillium wilt is the most effective and feasible way to combat the problem. Little is known about the inheritance of resistance to Verticillium wilt of cotton, especially that caused by the defoliating (D) and nondefoliating (ND) pathotypes of the soil‐borne fungus V. dahliae. The objective of this study was to determine the inheritance of resistance in cotton against both pathotypes of V. dahliae. Crosses were made between the susceptible parent ‘Cukurova 1518’ and each of four resistant parents PAUM 401, PAUM 403, PAUM 405 and PAUM 406 to produce F₂ generations in 2002 and F_2:3 families in 2003. Disease responses of parent and progeny populations to the D and ND pathotypes were scored based on a scale of 0‐4 (0, resistant; 4, susceptible). F₂ populations inoculated with the D pathotype showed a 3 : 1 (resistant : susceptible) plant segregation ratio. Tests of F_2:3 families confirmed that resistance was controlled by a single dominant gene. In contrast, analysis of data from F₂‐ and F₂‐derived F₃ families suggested that resistance to the ND pathotype is controlled by dominant alleles at two loci.     

DNA fingerprinting studies in coloured cotton genotypes

A collection of 11 coloured cotton Gossypium hirsutum genotypes and four white linted genotypes of different origin were evaluated by randomly amplified polymorphic DNA (RAPD) analysis. These 15 cotton genotypes were evaluated using 32 different 10‐mer primers of arbitrary sequences. All 32 primers were polymorphic ‐ in total, 287 amplified fragments were observed in these patterns. Out of the 287 fragments, 219 were polymorphic accounting for 76.31% of the total number of fragments. Similarity indices were calculated using the Dice coefficient and a dendrogram showing relationships between genotypes was obtained by Unweighted Pair Group Method of Arithmetic Average (UPGMA) cluster analysis. Cluster analysis showed clear‐cut separation of the coloured and white linted genotypes and thus formed three clusters (I, II and III). Among the coloured linted genotypes, all except ‘Parbhani American’ and ‘Lousiana brown’ clustered together. Cluster II contained white linted genotypes orginating from the same breeding station. The results indicate that RAPDs may constitute a relatively simple and efficient method for analysing genetic variation in coloured and white linted G. hirsutum collections.     

对一个新发现的棉纤维突变体的鉴定及特性分析

在农杆菌介导的转基因组织培养再生后代中发现了一个无绒有絮的纤维发育突变体,通过自交选择T₃代获得其纯合体,命名为CM突变体。尽管CM突变体从转基因后代中发现,但和转基因插入位点无关,推测是在组织培养过程中产生的点突变所致。通过与陆地棉遗传标准系TM-1,海岛棉军海1号,以及新乡小吉无绒有絮(XinFLM),新乡小吉无绒无絮(XinWX),徐州142无绒无絮(XZ142WX),显性光子N1N1,隐性光子n2n2、SL-7-1、MD17及T586等一系列纤维发育突变体分别配制F₂组合进行突变基因的遗传及等位性分析,结果表明CM突变体与纤维发育正常的TM-1和军海1号杂交,F₁表型均为无绒有絮,F₂表现无绒有絮和有绒有絮3∶1分离,说明该突变体与纤维发育正常材料相比,在短绒发育方面存在一个位点的差异,该突变性状由单显性基因控制。等位性测验及分子定位均表明, 控制该突变体短绒的基因与控制N1N1显性光子的N1基因等位。扫描电镜进一步证明该基因突变会造成纤维起始突起延迟。与N1N1突变体相比,CM突变体的衣分比N1N1显著高,而百粒重比N1N1极显著低。推测CM突变体中的突变基因与显性N1基因为复等位基因。     

转基因抗虫棉产量相关性状QTL的分子标记及定位

 采用亚洲棉渐渗的纤维强度突出的陆地棉优质新品系0-153与陆地棉转基因抗虫新品系sGK9708为亲本,构建了F₂及F_2∶3分离群体。利用3869对SSR引物筛选亲本,得到125对多态性引物。进一步对183个F₂群体单株分析得到150个多态性标记位点,其中100个标记位点连锁,构建20个连锁群,共覆盖660 cM,占棉花总基因组的14.67%,每个连锁群平均包含5个标记位点,标记间平均相距6.6 cM,其中13个连锁群确定了对应的染色体。利用F₂和F_2:3数据,通过复合区间作图,共检测到28个产量及相关因素的QTLs。这些控制产量性状的QTLs只存在于5个连锁群上,成簇分布。与皮棉产量性状有关的2个QTLs,均与其它多个产量相关性状的QTLs在同一个连锁区段内,增效基因遗传效应方向一致,有必要研究其在标记辅助选择中的效果。本研究没有检测到在多世代表现稳定的QTL。因此,需要培育重组自交系,进一步明确产量性状有关QTL的遗传效应。     

Molecular mapping of a new red mutant gene (Rs) in upland cotton

In recent years, there has been slow progress in improving cotton yield. It is known that the F₁ generation from the cross of the new red mutant and the normal green leaf plant has high photosynthetic efficiency. Therefore, cloning the new red mutant gene and further introducing it into other crops through transgenic techniques is a promising approach for achieving high photosynthetic efficiency through breeding. To map this new mutant gene, tentatively named R _s , the authors constructed an F₂ generation containing 1214 individual plants from mutant EH083 ( Gossypium hirsutum ) and Hai 7124 ( Gossypium barbadense ). Fifty-five pairs of simple sequence repeats and sequence-related amplified polymorphism (SRAP) primers on chromosome 7 were selected to screen the two parents. Finally, the R _s gene was mapped at the 0.3 cM interval flanked by markers NAU3735 and NAU1048.     

Assessment of cytoplasmic effects of cytoplasmic male-sterile lines in upland cotton

Paired A- and B-lines, cytoplasmic male-sterile (CMS)-lines and main-tainer lines, respectively, were crossed with R-lines (restorer) to produce A × R, R × B and B × R hybrids, which were used in 1993 and 1996 to predict the performance of three lypes of CMS cotton lines available in China. A significant difference in yield and yield components was revealed between paired A × R and B × R hybrids. This difference was greatly influenced by both CMS cytoplasm and the interaction between cytoplasm and nuclear genotypes. It is suggested that there are detrimental effects of CMS cytoplasm on yield and yield components. General combining ability of near-isogenic CMS lines was also affected by this negative effect. The detrimental effect was closely related to an increased number of immature seeds per boll, which might be caused by partial female sterility associated with CMS cytoplasm. The possibility of developing specific combinations of CMS Upland × Upland restorer hybrids that express enough heterosis for yield to overcome the detrimental effect of the CMS cytoplasm is discussed.     

The integration and insertion site of GbVe1 gene in the genome of transgenic cotton (Gossypium hirsutum)

[Objective] The aim of this study was to obtain the flanking sequences of T-DNA in the transgenic cotton containing a GbVe1 over-expression cassette. [Method] The T-DNA insertion copy number in the transgenic GbVe1 cotton was analyzed by southern blot. Flanking sequences of the transgenic lines with putative single T-DNA insertion copy were obtained using high-efficiency Thermal asymmetric interlaced polymerase chain reaction (hiTAIL-PCR). The T-DNA insertion sites were further confirmed by PCR with specific primers. [Result] RB-flanking sequences (119-1 018 bp) and LB-flanking sequences (243-516 bp) were obtained from three transgenic lines with low copy number of T-DNA insertion. The AT content was more than 63% in these flanking sequences. A same single insertion site in the intron of Gohir.D01G157600.1 was found in the two transgenic lines 7/100826-152 and 12/100826-393, while two separated insertion sites, one also in the intron of Gohir.-D01G157600.1 and the other in the intergenic region of A12 chromosome, were found in the transgenic line 1/w-ch14. A deletion of 21 bp was found in the insertion site in the intron of Gohir.D01G157600.1. The T-DNA insertion in the intron of Gohir.D01G157600.1 was further confirmed by the specific PCR. [Conclusion] The flanking sequences of T-DNA in the transgenic GbVe1 cotton were obtained and the specific transformation event in the intron of Gohir.D01G157600.1 was further confirmed by PCR.     

玉米光敏色素A1基因(ZmPHYA1)在棉花中的转化及分子鉴定

为评价玉米ZmPHYA1基因在棉花种质资源改良中的价值,本研究利用农杆菌介导法在陆地棉(Gossypium hirsutum)R15材料中进行了玉米ZmPHYA1基因的遗传转化。经过愈伤组织诱导、抗性愈伤筛选、体细胞分化诱导后获得棉花转基因再生植株。通过田间草铵膦除草剂筛选鉴定抗性植株,并利用PCR扩增其草铵膦抗性基因和目的基因ZmPHYA1进行分子鉴定,发现阳性植株对草铵膦除草剂具较好抗性,并可扩增到256 bp的草铵膦抗性基因和217 bp ZmPHYA1基因的特异条带。进一步通过免疫印迹检测表明,3个不同转基因株系中外源ZmPHYA1基因可正常表达约170 kD大小的蛋白,且在不同组织中该外源蛋白均可正常表达。此外,对转基因植株的不同农艺性状分析表明,转基因株系株高明显低于受体对照,而铃重和纤维长度等性状无明显差异。本研究成功获得具有草铵膦抗性和外源ZmPHYA1基因的棉花新种质材料,为进一步利用光敏色素基因创新种质资源提供了材料来源。     

ET-ISJ标记的开发及陆地棉遗传图谱构建

根据植物结构基因外显子拼接位点的保守序列,设计扩增外显子的ET-ISJ (exon targeted intron-exon splice junction)标记引物。利用1 280对ET-ISJ引物组合,在陆地棉品种渝棉1号和T586中,筛选获得69对多态性引物组合,占引物组合的5.4%。用多态性ET-ISJ引物组合检测(渝棉1号×T586)F_2:7重组近交系群体,得到70个位点。以70个ET-ISJ标记位点与523个SSR、59个IT-ISJ、29个SRAP和8个形态标记进行连锁分析,构建的遗传连锁图谱包括59个连锁群和673个位点(68个ET-ISJ、510个SSR、58个IT-ISJ、29个SRAP和8个形态标记)。连锁图覆盖3 216.7 cM,占棉花基因组的72.3%,标记间平均长度为4.8 cM。68个ET-ISJ标记分布于20条染色体。研究表明ET-ISJ标记多态性较高、稳定性好,可有效用于棉花与其他植物遗传连锁图谱构建。     

棉花转录因子GhWRKY4基因的克隆及特征分析

 本研究从陆地棉中克隆得到GhWRKY4基因,全长cDNA是1281 bp,包含1083 bp的开放阅读框,编码的360个氨基酸属于IIcWRKY转录因子家族。分析GhWRKY4基因结构表明其有三个外显子和两个内含子。GhWRKY4蛋白的亚细胞定位实验表明其定位在洋葱表皮细胞核。实时荧光定量分析结果显示GhWRKY4在棉花的根、茎、叶和花中均有表达。染色体步移得到了GhWRKY4的5’端侧翼序列,生物信息学软件预测表明GhWRKY4包含一系列和胁迫基因调控相关的反式作用元件,表明GhWRKY4参与植物对盐害和冷害胁迫的反应。