棉花品种遗传纯度的SSR分子标记鉴定技术研究

为建立适合于SSR标记的棉花品种纯度鉴定方法,利用78个SSR标记对12个棉花常规品种进行标记基因型分析.通过比较不同品种不同单株标记基因型,发现棉花品种的非纯SSR位点存在3种主要类型.通过综合分析非纯SSR位点率和异型单株率对品种遗传纯度的影响,建立了利用SSR标记在分子水平上鉴定棉花品种纯度的方法.利用该方法鉴定的12个棉花品种中有3个品种(C5、C9和Cll)的遗传纯度在98％以上,非纯SSR位点和异型株均较高的2个品种(C3和C6)遗传纯度分别为67.31％和31.79％,该方法弥补了以往分子纯度计算方法中只考虑单株混杂、不考虑SSR位点混杂的缺陷,可比较客观地反映品种的遗传纯度状况.     

The Effectiveness of HA-GGE Biplot Application in Analyzing the Data from the National Cotton Cultivar Regional Trial in the Yangtze River Valley

A large number of regional crop trials have demonstrated the ubiquitous existence of genotype × environment interactions (G×E), which make it complicated to select superior cultivars and identify the ideal testing sites. The GGE (genotype main effect plus genotype × environment interaction) biplot is the most powerful statistical and graphical displaying tool available for regional crop trial dataset analysis. The objective of the present study was to demonstrate the effectiveness of the biplot in evaluating the high and stable yields of candidate cultivars simultaneously, and in delineating the most adaptive planting region, analyzing trial location discrimination ability and representativeness, and identifying the ideal cultivar and trial locations. The lint cotton yield dataset with nine experimental genotypes and 17 test locations (three replicates in each) was collected from the national cotton regional trial in the Yangtze River Valley (YaRV) in 2012. The results showed that: (1) the effects of genotype (G), environment (E), and genotype × environment interaction (G×E) were significant (P < 0.01) for lint cotton yield. Differences among environments accounted for 78.7% of the treatment total variation in the sum of squares, whereas the genotype main effect accounted for 8.7%, and the genotype × environment interaction accounted for 12.6%. (2) The “ideal cultivar” and “ideal location” view of the HA-GGE biplot identified Zhongcj408 (G2) and Nannon12 (G9) as the best ideal genotypes; Cixi in Zhejiang Province and Jiangling in Hubei Province were the most ideal locations.(3) The “which-won-where” view of the biplot outlined the adaptive planting region for each experimental cultivar. (4) The “similarity among locations” view clustered the trial locations into four groups, among of which the two outlier locations, Shehong (SH) and Chengdu (QBJ), located in Shichuan Basin in the upper reaches of YaRV, were clustered in one group, whereas the Nanyang (NY) of Henan Province at the northern edge of YaRV was singled out as a sole group. Such location clustering results implied an apparent association with the geographical environment.     

Leaf Temperature and Transpiration of Field Grown Cotton and Soybean under Arid and Humid Conditions

Abstract

Abstract : Cotton (Gossypium hirsutum L.) and soybean (Glycine max (L.) Merr.) cultivars were grown under arid (Urumqi, Xinjiang, China) and humid (Matsudo, Chiba, Japan) conditions to analyze their abilities to adapt to arid conditions in terms of transpiration, leaf movement and leaf temperature. Under the arid condition, the leaf temperature of the cotton cultivars was higher than that of the soybean cultivar and the air temperature. There was no significant difference in leaf temperature among the cotton and soybean cultivars under the humid condition. The flow rate of stem sap in the cotton cultivars under the arid condition was always higher than that in the soybean cultivar, and was largely affected by vapor pressure deficit (VPD). Under the humid condition, however, the flow rates of stem sap were lower in the cotton cultivars than in the soybean cultivars. These results indicate that cotton can avoid heat stress by the high transpiring ability possibly supported by well-developed root systems, which leads to higher drought resistance under the arid condition. Soybean would adapt to arid conditions by the combination of paraheliotropic leaf movement and reduced transpiration.     

Agronomic traits at the seedling stage,yield, and fiber quality in two cotton (Gossypium hirsutum L.) cultivars in response to phosphorus deficiency

ABSTRACT

Cotton is critical for phosphorus demands and very sensitive for its deficiency. However, identifying the effect of low-phosphorus tolerance on cotton growth, yield, and fiber quality by reducing phosphorus consumption. This may help to develop phosphorus-tolerant high-yielding cotton cultivars. In a two-year repeated (2015 and 2016) hydroponic experiment (using 0.01 and 1 mM KH₂PO₄), two cotton cultivars with phosphorus sensitivity (Lu 54; a low-phosphorus sensitive and Yuzaomian 9110; a low-phosphorus tolerant) were screened on the base of agronomic traits and physiological indices through correlation analysis, cluster analysis and principal component analysis from 16 cotton cultivars. Low phosphorus nutrition reduced the plant height, leaf number, leaf area, phosphorus accumulation and biomass in various organs of seedlings. The deficiency negatively affected the morphogenesis of seedlings, as well as yield and fiber quality. Further, these screened cultivars were tested in a pot experiment with 0, 50, 100, 150, 200 kg P₂O₅ ha^?1 during 2016 and 2017. It was found to have a significant (P< 0.05) difference in boll number, lint yield, fiber strength, and micronaire at the harvest. Furthermore, after collectively analyzed the characteristics of Lu 54 and Yuzaomian 9110, there were six key indices that could improve the low phosphorus tolerance of cotton cultivars. These were root phosphorus accumulation, stem phosphorus accumulation percentage, leaf and total biomass of seedlings, seed cotton weight per boll and fiber length.     

Isolation and Characterization of GhLEA3 Gene from Upland Cotton and Its Expression in Response to Low Temperature Stress

[Objective] Based on the analysis of cotton GhLEA3 gene structure and its expression model in chilling stress, we investigated the resistance function of GhLEA3 in response to cold stress. [Method] We cloned GhLEA3 from upland cotton by homologous sequence cloning way. We analyzed the properties of the GhLEA3, and constructed phylogenetic tree by bioinformatics analysis. We constructed the transient expression vector 35S∷GhLEA3-GFP by In-Fusion connection technology and studied the subcellular localization of GhLEA3. The expression of GhLEA3 gene in leaves of TM-1 at three-leaf stage under low temperature stress treatments (4 ℃, 24 h) was performed. Agrobacterium tumefaciens mediated floral dipping method was used to transform the gene with expression vector 35S∷GhLEA3-GFP into Arabidopsis thaliana wild type. Low temperature germination experiment at 4 ℃ was conducted to verify germination ability of the transgenic Arabidopsis thaliana T₃ seed. The transgenic Arabidopsis seedlings were treated at 4 ℃(low temperature), and all the leaves were taken to measure their electrical conductivity. [Result] The coding sequence of GhLEA3 gene is 1 218 bp, which encodes 405 amino acids. GhLEA3 protein includes 4 functional domains of PF02987. Phylogenetic analysis showed that the similarity of amino acid sequences between upland cotton GhLEA3 and Arabidopsis thaliana AtLEA3 was 52.6%. GhLEA3 might be mainly localized in vacuoles and small vesicles. GhLEA3 was up-regulated in leaves after low temperature treatment. The germination rate of transgenic Arabidopsis thaliana of T₃ generation at low temperature was significantly higher than that of wild type, and its electrical conductivity of leaves after low temperature treatment was significantly lower than that of wild type. [Conclusion] GhLEA3 belonged to the member of LEA3 family. Phylogenetic analysis showed that GhLEA3 had the closest relationship with AtLEA3. GhLEA3 was induced by low temperature stress and may play an important role in improving cold resistance.     

Identification of resistance to Aspergillus flavus infection in cotton germplasm

Natural resistance in cottonseed to Aspergillus flavus infection has not been explored to date. A green fluorescent protein (GFP) expressing A. flavus strain was used to assess the resistance of seed from 35 cotton varieties from Gossypium arboreum, G. barbadense, and G. hirsutum. Mature cotyledons devoid of seed coat were wounded, inoculated, and assessed for innate resistance to A. flavus infection. Of the initial 35 varieties tested, we observed a range of resistance to infection in representatives of all three species. A subset of 15 representatives was further analyzed. Within this group, G. arboreum cultivar A2 186 and G. hirsutum cultivar SA 1582 were most resistant to fungal infection. The most susceptible cultivar in this group was G. hirsutum SA 1595. The remaining 12 representatives tested in the secondary screen (3 G. arboreum, 3 G. barbadense, and 6 G. hirsutum lines) exhibited intermediate resistance. We did not observe any relationship between species and resistance. Within each species, there was a range of responses to fungal infection. Future studies using this methodology to screen additional diploid and tetraploid cotton lines may enable us to identify naturally resistant germplasm that can be used to develop cotton with enhanced resistance to fungal infection.     

P0 protein of cotton leafroll dwarf virus-atypical isolate is a weak RNA silencing suppressor and the avirulence determinant that breaks the cotton Cbd gene-based resistance

Cotton blue disease (CBD) is the most important disease present in cotton crops in South America and cotton leafroll dwarf virus (CLRDV) is the causal agent. The disease has been controlled by sowing cotton varieties resistant to CLRDV. However, in the 2009/10 growing season, an outbreak due to an atypical CLRDV isolate (CLRDV-at) occurred in northwest Argentina. Although CLRDV and CLRDV-at genomes are very closely related, the symptoms they produce in cotton plants are quite different. P0 is the most divergent protein between the isolates and in CLRDV is a silencing suppressor protein. This work characterized the silencing suppressor activity of the P0 protein encoded by CLRDV-at (P0^CL-at) and evaluated its role in Cbd-resistance break in cotton plants. It was demonstrated that P0^CL-at, despite having a mutation in the consensus of the F-box-like motif, was able to suppress local RNA silencing, but displayed lower activity than P0^CL. P0^CL and P0^CL-at showed no differences in the interaction with Gossypium hirsutum SKP1 orthologue (GSK1) and Nicotiana benthamiana SKP1 and both P0 proteins triggered destabilization of ARGONAUTE1. However, when the ability to enhance PVX symptoms was evaluated, P0^CL-at was shown to be a weaker pathogenicity factor than P0^CL in N. benthamiana. Interestingly, trans-expressed P0^CL-at enabled CLRDV to systemically infect CBD-resistant plants, and a chimeric CLRDV-P0^CL-at infectious clone succeeded in establishing infection in CBD-resistant cotton varieties with symptoms resembling those produced by CLRDV-at. These results strongly suggest that P0^CL-at is the avirulence (Avr) determinant involved in breaking cotton Cbd gene-based resistance.     

陆地棉原生质体培养与植株再生

以陆地棉品种“珂字２０１”为材料,比较了ＩＡＡ＋ＫＴ和２,４－Ｄ＋ＫＴ在愈伤诱导和悬浮培养中的效应,结果表明：愈伤组织诱导中,ＩＡＡ和２,４－Ｄ表现为正效应,且２,４－Ｄ的效应强于ＩＡＡ;ＫＴ表现为负效应;胚性愈人务悬浮培养中;３种激素都表现出负效应。以胚性细胞悬浮系了原生质体的分离和培养试验,分离原生质体的最佳酶组合为纤维素酶３％＋果胶酶１．５％,原生质体培养的最佳激素为ＩＡＡ０．５ｍｇ／Ｌ＋Ｋ