An extended random primer amplified region (ERPAR) marker linked to a dominant male sterility gene in cabbage (Brassica oleracea var. capitata)

Similar to SCAR, an extended random primer amplified region (ERPAR) marker is a PCR amplified genomic DNA fragment at a single genetically defined locus. However, ERPAR uses specific primer pairs derived from RAPD primers by adding bases sequentially to their 3′-ends. As an example, an ERPAR marker was derived from a RAPD marker (OT11₉₀₀) linked to a dominant male sterility gene in cabbage (Brassica oleracea var. capitata). After two cycles of base adding and primer pair screening, a primer pair (5′-TTCCCCGCGACT-3′and 5′-TTCCCCGCGAGA-3′) amplified a single intense band with the same size as OT11₉₀₀. The identity of the new marker and OT11₉₀₀ was verified by segregation analysis. The new marker amplified by this extended primer pair was named as EPT11₉₀₀. The development of ERPAR exploits the importance of 3′-end bases of primers in PCR ERPAR shares advantages of SCAR, but eliminates the need for cloning and sequencing. It is a fast and universal way of converting RAPD markers into stable markers. This revised version was published online in July 2006 with corrections to the Cover Date.     

结球甘蓝的离体快繁技术研究

以结球甘蓝无菌苗的胚轴和子叶为材料，几乎不生长愈伤组织，分化出不定芽。不定芽在添加6-BA的MS培养基上增殖很快，适宜的生根培养基为1/2MS+IBA0.5mg/L+ NAA0.1mg/L。通过增加培养基中糖和琼脂粉的用量，提高培养时的光照强度等有效控制了玻璃苗的产生。     

Identification of a Monogenic Dominant Male Sterility Mutant in Broccoli (Brassica oleracea var. italica Plenck)

After mutagenic treatment with NMU (nitroso-methylurea) flower buds and flower-bud pedicels of broccoli were cultivated in vitro. Among regenerated MJ plants, one male sterile, but female fertile mutant was found, for which a monogenic dominant inheritance of male sterility was demonstrated. A scheme to utilize dominant controlled genie male sterility in Brassica oleracea for hybrid breeding is discussed.     

Bud pollination and hybrid seed production in detergent-induced male sterile plants of Brassica juncea

The efficacy of a synthetic detergent (Surf Excel) as a potential chemical hybridizing agent in Brassica juncea was studied. Foliar sprays with various concentrations of the detergent caused reductions in plant height, number of branches and leaves per plant, size of leaves, anther size, pollen per flower, ovules per flower, pollen fertility, fruits per plant, fruit size, seeds per fruit, total yield per plant and 100 seed weight as compared with those of untreated plants. The style in the floral buds of plants sprayed with different concentrations of Surf Excel elongated and so the receptive stigma protruded from the buds which facilitated cross‐pollination by honey bees. The plants sprayed once with 2% Surf Excel exhibited an elongated style with a raised receptive stigma and 100% pollen sterility without causing a significant reduction in total yield.     

Molecular mapping of a dominant genic male sterility gene Ms in rapeseed (Brassica napus)

Rs1046AB is a genic male sterile two‐type line in rapeseed that has great potential for hybrid seed production. The sterility of this line is conditioned by the interaction of two genes, i.e. the dominant genic male sterility gene (Ms) and the suppressor gene (Rf). The present study was undertaken to identify DNA markers for the Ms locus in a BC₁ population developed from a cross between a male‐sterile plant in Rs1046AB and the fertile canola‐type cultivar ‘Samourai’. Bulked segregant analysis was performed using the amplified fragment length polymorphism (AFLP) methodology. From the survey of 480 AFLP primer combinations, five AFLP markers (P10M13₃₅₀, P13M8₄₀₀, P6M6₄₁₀, E7M1₂₃₀ and E3M15₁₀₀) tightly linked to the target gene were identified. Two of them, E3M15₁₀₀ and P6M6₄₁₀, located the closest, at either side of Ms at a distance of 3.7 and 5.9 cM, respectively. The Ms locus was subsequently mapped on linkage group LG10 in the map developed in this laboratory, adding two additional markers weakly linked to it. This suite of markers will be valuable in designing a marker‐assisted genic male sterility three‐line breeding programme.     

结球甘蓝(Brassica oleracea var. capitata)迟抽薹基因SCAR标记转CAPS标记

为了更加精确结球甘蓝迟抽薹基因标记。本研究以结球甘蓝冬性强的迟抽薹材料‘P02’和冬性弱的易抽薹材料‘A21’杂交得F1代,F1代自交至F3代为试验材料,并以课题组所设计的特定序列扩增(sequence characterized amplified regions, SCAR)标记的SCN1/248为引物。在F3代的785株结球甘蓝中有711株扩增出目的条带,有74株未能扩增出目的条带。经测序,该序列长度为248 bp,运用该序列共设计3对酶切扩增多态性序列(cleaved amplified polymorphic sequence, CAPS)引物,其中1对引物均扩增出目的条带,多态性消失,另外两对分别命名为SC01和SC02。结合田间127株结球甘蓝抽薹性状调查,有65株表现为迟抽薹,59株表现为易抽薹,3株表现为中间型,与SCAR扩增检测结果基本一致,将SCAR标记转换成CAPS标记,建立CAPS标记体系。本研究进一步精化了分子标记辅助选择,为做精确定位功能基因,选育优良的迟抽薹结球甘蓝品种提供了理论依据。     

Screening and Identifying Two Specific Molecular Markers in Maintainer Line of Cytoplasmic Male Sterility Cauliflower （Brassica oleracea var. botrytis）

Cytoplasmic male sterility （CMS） is a maternally inherited trait that prevents the production of function pollen, but maintains female fertility. It has been widely used in breeding programs to product F1 hybrid seed in some crops （Hanson, 1991）. However, in some plants the CMS lines usually have some harmful characters such as plant dwarfness, etiolation in cauliflower. To obtain perfect CMS lines, a long time usually five to eight years is necessary. So how to fully utilize the obtained CMS lines for accelerating the development of new hybrid seed is very important. Here, in order to select high-quality paternal materials for cytoplasmic male sterility （CMS） cauliflower NKC-A in early stage, analyses of RAPD and ISSR were performed. Total 2 160 detectable bands were obtained by RAPD using 406 random primers. Among all these primers only the amplifications of primer S2121（5＇ GTGGCTCTCC 3＇） were polymorphic. A 934 bp band only existed in maintainer line. After cloning and sequencing, specific primers were designed to transform the RAPD marker into specific PCR marker, which was named S2121900. Furthermore, total of 30 ISSR primers were also analyzed which produced 306 detectable bands.     

春甘蓝抽薹的生理与相关基因转录分析

本研究以高代自交获得的4份具有不同抽薹时间且遗传稳定的春甘蓝品系为材料,在观察其园艺学性状的基础上,分析了抽薹性状与可溶性蛋白和可溶性糖含量之间的关系,结果显示,耐抽薹春甘蓝品系的可溶性蛋白和可溶性糖含量低于不耐抽薹的,抽薹前春甘蓝的可溶性蛋白和可溶性糖含量也均低于抽薹后的.进一步的转录分析结果表明,5个与植物抽薹开花...     

Inheritance of Waxlessness in Kohlrabi (Brassica oleracea L. var. gongyloides) and its Utilization in Hybrid Seed Production

By genetic analysis of selfed progenies and reciprocal F₁, F₂ and B₁: populations of crosses of a waxless mutant and three waxy plants two recessive genes were identified in kohlrabi (Brassica oleracea L. var. gongyloides), causing waxlessness independently from each other. Gene a is responsible for glossy waxless and gene b tor dull waxless plants. To be able to form a wax layer a plant must have both genes in the dominant constitution, i.e. A.B. (“complementary gene effect). Some segregation results which deviate from expectation could indicate a potential linkage between one of the two wax loci and the incompatibility locus. Both genes can be used as markers m hybrid breeding of cole crops as well as in basic research.     

Chromosomal monogenic dominant male sterility in chinese cabbage (Brassica rapa supbs. pekinensis (Lour.) hanelt)

Summary Strong indication was found for the existence of a chromosal monogenic dominant male sterility-gene in chenese cabbage. This source of male sterility can be of practical use for the production of hybrid varieties. A pronounced drawback is the required removal, in the breeding and seed production stage, of the approximately 50% male fertile plants from each offspring of male sterile plants. Fortunately this removal is facilitated by the penomenon of apical dominace in the flowering process in the whole Brassica rapa taxon. Moreover an advantage of dominant monogenic male sterility is its easy and rapid introduction into all available genotypes of chinese cabbage.     

甘蓝细胞质雄性不育相关基因orf138的分子特性分析

为了研究甘蓝雄性不育机制,根据萝卜CMS相关基因orf138的序列信息,设计特异引物,并在甘蓝不同类型不育系和保持系中鉴定PCR产物的稳定性。随后利用Tail-PCR技术,扩增获得此基因的侧翼序列并进行了生物信息学分析。结果表明在甘蓝不育型材料中,能够稳定扩增出300 bp左右的单一条带,而在其他细胞质不育类型和可育材料中均未扩出条带,经多次验证结果稳定可靠。甘蓝中orf138的上下游侧翼序列有效碱基1789 bp,通过生物信息学分析,获得包括起始密码和终止密码的orf138的完整序列共417 bp。同源性比对结果显示：与甘蓝型油菜﹑白菜和萝卜的orf138片段具有高度保守性。分析侧翼序列表明甘蓝orf138的3’端是由ORF83、trnfM、ORF125等基因片段构成的一个复杂序列。获得了甘蓝OguCMS特异的分子鉴定标记,明确了orf138在甘蓝线粒体中的位置,以上结果为甘蓝雄性不育的进一步研究奠定良好的基础。     

The allelic relationship between A IP(internal pigmentation) and A RC (red cabbage) in Brassica oleracea var. capitata L.

Summary Internal anthocyanin pigmentation (IP), previously reported to be controlled by a single gene dominant to aa of normal green cabbage, was studied further to determine its relationship to A ^RC A ^RC of red cabbage. When IP line R52 was crossed with an early red cabbage line. F₁ heads were pigmented throughout, but pigment intensity was intermediate. Subjective classification of F₂ plants by pigmentation intensity and distribution scores gave a ratio of 1 intense red throughout (red cabbage):2 medium red throughout:1 medium to light red, restricted to the central portion of the head (IP). The genotypes A ^RC A ^RC: A ^RC A ^IP: A ^IP A ^IP. respectively, are proposed to explain these 3 phenotypic classes. F₂ progenies contained no normal green plants, supporting the conclusion that A ^IP and A ^RC are alleles.Technical Paper 7052, Oregon Agricultural Experiment Station.     

Functional male sterility (ps-2) in tomato (Lycopesicon esculentum Mill.) and its application in breeding and hybrid seed production

Data and findings concerning the development of an effective male sterility system (as already applied in the practice of producing tomato hybrid seed) through the use of the positional sterility (ps 2) gene are reported. Undesirable selfing and the necessity of emasculation are discussed as the two main disadvantages that limit the use of ps 2-sterile seed parents in tomato hybrid seed production. The following specific characteristics in the performance of ps 2-lines were evaluated: 1) the percentage of selfing in the ps 2-lines varies within and between the years of growing, being forever lowest during the period of hybrid seed production; 2) the percentage of selfed seeds when using a ps 2-line as seed parent in producing hybrid seed is significantly lower than the percentage of selfing, observed on this same line; 3) the ps 2-gene expressivity varies depending on the genotype. This enables the breeding of ps 2-lines exhibiting very low percentage of selfing; 4) stamen emasculation at anthesis (as applied in ps 2-lines) is significantly easier and more rapid than emasculation in floral buds. The easy maintenance of the sterile lines – by artificial selfing it is possible to produce 100% ps 2 progeny – and the high yield of hybrid seed obtained are discussed as important advantages in using this type of sterility in tomato hybrid seed production. This revised version was published online in July 2006 with corrections to the Cover Date.     

The somatic karyotype of cabbage (Brassica oleracea ssp. capitata)

Summary The individual chromosomes of cabbage are identifiable in nucellar cells found in the ovaries of immature pistils by the combination of their relative length, arm ratios, secondary constrictions, chromatic and achromatic regions and the presence of satellites.     

An S-allele survey of cabbage (Brassica oleracea var. Capitata)

Summary A total of 31 S-alleles was found in a survey of 197 cabbage plants representing 11 cultivars of diverse type. Most of these S-alleles also occurred in either kale or Brussels sprouts, but five of them have not been found previously and apparently occur only in cabbage. A more detailed study of five cultivars of spring cabbage showed only 12 S-alleles in all, with 6–10 S-alleles in four older cultivars and only 3 S-alleles in the newer more highly selected cultivar. S2 was by far the commonest S-allele, as it is in B. oleracea as a whole. The highly recessive alleles S5 and S15 were not particularly common in cabbage and this may partly explain why the sib problem in F₁ hybrids is apparently less in cabbage than in Brussels sprouts. Three cases were found in which an S-allele was completely recessive in both the stigma and the pollen. The problems for the breeder created by this rather unusual situation are discussed.     

Plant traits associated with resistance to Thrips tabaci in cabbage (Brassica oleracea var capitata)

Thrips tabaci is a major problem in the cultivation of cabbage for storage, as this pest causes symptoms that necessitate the removal of affected leaves from the product. Between cabbage varieties large differences in susceptibility occur. This study aimed to identify plant traits associated with these differences, in field experiments with natural infestation in 2005 and 2006. One factor affecting the amount of thrips damage was the timing of the development of the head. In an experiment with different planting dates especially the early maturing, more susceptible varieties were shown to benefit from later planting. In comparisons of multiple varieties in both years, regression studies showed that more advanced plant development in August and early September increased thrips damage at the final harvest. However, no single plant trait explained more than 25% (2005, Brix) or 48% (2006, compactness) of the variation in thrips damage. Optimal regression models, explaining up to 75% of the variation in thrips damage included Brix and leaf surface wax late in the season, as well as an indicator of plant development earlier in the season, and in 2005 also leaf thickness. The possible role of these plant traits in relation to thrips is discussed.     

自花授粉诱导的甘蓝功能基因BoSPI的克隆与表达分析

自交不亲和性是甘蓝在长期进化过程中形成的防止自交衰败、促进杂交优势的一种复杂而完善的遗传机制。克隆自交不亲和性相关基因对甘蓝自交不亲和性的深入研究和利用有重要意义。本研究通过挖掘0~60 min自花和异花授粉的甘蓝柱头转录组数据, 筛选到一个受自花授粉诱导上调表达的基因, 命名为BoSPI。BoSPI开放阅读框534 bp, 编码177个氨基酸, 理论等电点为4.21, 不包含信号肽和跨膜区, 含有4个保守的EF-hand结构域。BoSPI基因起始密码子上游2000 bp的核苷酸序列中含有真菌诱导响应、代谢调节以及器官形成等应答元件。BoSPI基因在大肠杆菌中可诱导表达为17 kD的蛋白。BoSPI在柱头中表达量最高, 在花瓣、萼片、叶片、雄蕊表达量较低。BoSPI蛋白被定位在细胞膜和细胞质。自花授粉30 min后对BoSPI基因的诱导表达显著增强。表明BoSPI参与了柱头响应自花花粉刺激的分子过程, 可能是实现甘蓝自交不亲和性相关的某种新功能基因。     

甘蓝型油菜白花突变体的遗传研究和白花胞质雄性不育系的选育

用白花突变体和白花不育突变体分别选育白花品系G95150和白花胞质不育系7600 A。遗传研究表明,白花对黄花由一对不完全显性基因控制。7600A在农艺性状上与其保持系G95150完全一致：叶色浓绿,植株较矮（163.5 cm）、生育期较长（243天）、抗寒能力较差（冻害率100%,冻害指数33.8）。7600A花色纯白、花瓣皱缩、花药小、低温存在微量花粉。G95150和7600 A的芥酸含量为0.8854 %,硫甙含量70.5461μmol/g。