甘蓝型油菜耐盐和耐旱相关性状的QTL分析

盐胁迫和干旱胁迫是非生物胁迫中影响作物产量的重要因素,检测与耐盐和耐旱相关的QTL,可为抗逆油菜品种的选育提供理论依据。本研究利用德国冬性甘蓝型油菜Express和中国半冬性甘蓝型油菜SWU07为亲本构建的包含261个株系的双单倍体(doubled haploid, DH)群体,分别以1.2%NaCl溶液和20%PEG-6000溶液作为培养液模拟盐胁迫和干旱胁迫,去离子水为对照,对2个亲本和DH群体进行发芽试验。播种后7 d测定幼苗根长、鲜重及发芽率,计算各性状在盐胁迫和干旱胁迫下的相对值,并作为评价耐盐和耐旱的指标。根据已构建的遗传连锁图谱进行QTL定位。盐胁迫下,在3次重复中共检测到与盐胁迫相关的QTL12个,分布在A02、A03、A05、A09、C01和C09染色体上,单个QTL可解释的表型变异为3.61%～10.59%,其中5个QTL在不同的重复中被检测到。干旱胁迫下,共检测到与干旱胁迫相关的QTL 9个,分布在A01、A02、A03、A05、A09、A10和C03染色体上,单个QTL可解释的表型变异为3.94%～12.90%,其中2个QTL在不同的重复中被检测到。此外,在A0...     

Occasional loss of expression of phosphinothricin tolerance in sexual offspring of transgenic oilseed rape (Brassica napus L.)

The commercial and economic value of genetically modified crops is determined by a predictable, consistent and stable transmission and expression of the transgenes in successive generations. No gene inactivation is expected after selfings or crosses with non-transformed plants of homozygous transgenic oilseed rape plants if the expression of the transgene in homozygous or hemizygous nature in such plants is stable. The segregation ratios of phosphinothricin (PPT) tolerance in successive generations of selfings and mutual crosses of a few independent transgenic PPT-tolerant oilseed rape plants indicated a dominant, monogenic inheritance. In within-variety and between-variety crosses no transgene inactivation was observed. However, after selfings and backcrosses with non-transgenic oilseed rape infrequent loss of the expression of the PPT tolerance transgene was observed independent from its homozygous or hemizygous nature. Molecular analysis of PPT-susceptible plants showed that the loss of expression was due to gene inactivation and not to the absence of the transgene. Methylation and co-suppression are mechanisms that might cause reduced or even loss of expression of the transgene in later generations. The implications of this observation for seed multiplication of varieties and breeding activities with transgenic oilseed rape are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Self-incompatibility as a pollination control mechanism for spring oilseed rape,Brassica napus L.

Summary Self-incompatibility was shown to be an effective method of pollination control in spring rapeseed (B. napus L. ssp. oleifera (Metzg.)) by comparing the yield of a Westar-Topas syn-1 produced by crossing two SI lines with the yield of the corresponding syn-1 produced by hand pollination. Although the trial showed high-parent heterosis in the syn-1s, there was insufficient replication to determine the level of heterosis.Abbreviations SI self-incompatible - SC self-compatible     

甘蓝型春油菜早花位点加密及位点聚合创建优异早花资源

cqDTFA7a and cqDTFC8, two major effects of early flowering QTL, were identified in the NNDH population of spring Brassica napus, and closely linked markers SSR G1803, InDel IA7-4, and SSR S035 with cqDTFC8 developed in previous studies. In this study, a BC₂F₂ population for early flowering QTL locus cqDTFC8 was constructed, and a closely linked SNP marker was further developed. The early flowering genotypes of one natural resources contained 93 spring B. napus varieties were identified used four closely linked markers with two loci, and selected 3164 and 2216 resources with cqDTFA7a site, 3484 and 2857 resources with cqDTFC8 site. Two site resources were aggregated by site polymerization through reciprocal hybridization. The polymerized DH system was rapidly obtained by microspore culture and maker assisted selection. A hybrid combination was created between polymeric line with good traits and early flowers and the Polima CMS, and the utilization value of the polymeric line was further analyzed by the production test at multiple environments for two consecutive years. cqDTFC8 encryption results showed that this site was located in the SNP11 and SNP12 interval, and separated from SNP11 altogether. The identification results of early flowering genotypes of natural resources showed that there were 50 individuals containing cqDTFA7a locus, with an average initial flowering period of 58.1 days; 16 single plants containing cqDTFC8 locus, with an average flowering period of 58.3 days; and 16 single plants containing two loci, with an average flowering period of 55.2 days, indicating the more early flowering sites containing, the earlier flowering. The results of polymerization showed that the flowering time of the polymerized lines of cqDTFC8 and cqDTFA7a was 2-3 days earlier than that of the single locus parents, among which the polymerized DH18 from 3164 of cqDTFA7a and 3484 of cqDTFC8 was 3 days earlier than that of the parents, and the yield-related traits were better than those of other lines. The combination of DH18 and the Polima CMS 025A was further utilized, and the combination was named TZG18. Yield results of two years and nine environments showed that yield of TZG18 was above 17.5% higher than the Haoyou 11, a local B. rapa variety on the Qinghai-Tibet plateau. Those results indicated that the early flowering site polymeric lines had an obvious advantage over the single locus lines in flowering time, and had an effect on the increasement of rapeseed yield. This study is a preliminary exploration of MAS breeding for early flowering traits of Brassica napus, providing materials support for replacing B.rapa varieties using early maturity Brassica napus varieties in spring rapeseed region, and approaches for gene polymerization breeding technology.     

Selection on pollen for linolenic acid content in rapeseed, Brassica napus L.

In order to enhance the economic value of edible rapeseed oil, an improvement of quality is necessary. Mutagenesis of rapeseed resulted in a low linolenic acid content and a low ‘linolenic acid (CIS: 3) level to linoleic acid (CIS: 2) level’ ratio, that is, the linoleic desaturation ratio (LDR), in the seeds of the Canadian variety ‘Stellar’. As an early breeding marker for low linolenic acid content, the pollen fatty acid composition was determined on 80 doubled haploid plants derived from a single F₁ hybrid obtained from a cross between ‘Stellar’ and a high CIS: 3 variety ‘Drakkar’. Fatty acid analysis on seed and pollen showed that the low CIS: 3 and the low LDR traits from the ‘Stellar’ variety were expressed in pollen and in seeds, and that a very close correlation (r = 0.88) existed between seed and pollen for these two traits. The inheritance of these traits is controlled by two major genes with additive effects, both in seed and pollen. However, minor genes also appeared to be expressed in pollen and seed. These genes may allow the production of plants with lower CIS: 3 levels than that of the low linolenic acid content parent. The efficiency of this new tool for early screening in breeding programmes is discussed.     

Abundance, polymorphism and genetic mapping of microsatellites in oilseed rape (Brassica napus L.)

The objective of the present study was to estimate the abundance and degree of polymorphism of simple sequence repeat (SSR) markers in rapeseed. By screening about 45000 clones of a small inserts library of rapeseed total DNA the abundances of GA/TC and CA/TG simple sequence repeats in the rapeseed genome were estimated to be approximately one repeat every 100 kb and 400 kb, respectively. After sequencing 13 positive clones, primer pairs could be designed for 11 microsatellite loci. Seven of these primer pairs produced reproducible amplification products in a set of 31 rapeseed genotypes, with one pair amplifying two independent products, giving a total of eight amplified loci. The different microsatellite loci displayed between one and three visible alleles. At four loci, additional null alleles were observed. With up to four alleles, polymorphic microsatellite markers show significantly higher allele numbers in rapeseed than restriction fragment length polymorphism (RFLP) markers. Four of the eight microsatellite markers could be mapped on four different linkage groups of an RFLP map of the rapeseed genome.     

Evaluating the effectiveness of isolation distances for field plots of oilseed rape (Brassica napus) using a herbicide-resistance transgene as a selectable marker

The effectiveness of 200- and 400-m isolation distances were evaluated for small-scale trials of oilseed rape (Brassica napus L.). These distances have been used for previous transgenic release experiments and are commonly adopted for production of basic and certified breeders seed. A 400-m² donor plot contained plants with a dominant transgene conferring resistance to the herbicide glufosinate-ammonium. This character was used as a selectable marker to measure the frequency of hybrid formation in 400-m² target plots of non-transgenic rape. Seeds produced by the non-transgenic plants were permitted to fall onto the plots and allowed to germinate. At the first true-leaf stage, the plants were sprayed with glufosinate-ammonium. Surviving plants were resprayed, and a subsample assayed for the presence of the transgene using a colorimetric assay and Southern-blot analysis. The average frequency of hybridization over two replicates was 0.0156% at 200 m and 0.0038% at 400 m. These estimates are within the limits established for the production of basic seed (0.1%). Results indicated that bees were the most likely agent for long-distance pollen dispersal.     

甘蓝型油菜小孢子培养胚发生能力的遗传分析

用 4× 4完全双列杂交研究了甘蓝型油菜小孢子培养胚状体发生能力的遗传规律。结果表明 :多数F1的胚产量与具高胚发生能力的亲本相近 ,部分F1的胚产量与双亲中亲值接近。胚状体发生能力主要由基因的加性效应控制。高胚发生能力由显性核基因控制。广义和狭义遗传力分别为 97 2 %和 81 1%。由Lisandra(高胚发生能 )×Kamikita(低胚发生能 )的F2 群体胚发生数的分离结果得出小孢子胚状体发生能力由具加性效应特点的两个基因位点控制     

甘蓝型油菜离体小孢子胚胎发生能力的遗传分析

对一套甘蓝型油菜6×6半双列杂交组合和1个F2群体进行了离体小孢子培养。 不同基因型(组合)间产胚率差异极显著, 小孢子产胚率的广义遗传力为0.85。 F1小孢子产胚率具有明显的杂种优势, 平均优势率为10.52%。 小孢子产胚率的一般配合力和特殊配合力方差均达到极显著。 根据一般配合力的大小, 可将供试亲本分为高和低配合力     

几个甘蓝型油菜品系主要农艺性状配合力和遗传力研究

弄清德国引进双低、高含油量材料作亲本的潜力,为科学地应用这几份材料提供理论基础,本研究以13份德国引进甘蓝型油菜作母本,与6个优良自交系进行不完全双列杂交,研究其在主要农艺性状上亲本间的配合力和性状的遗传力,结果表明：（1）一般配合力上,6个父本的优劣顺序为：042269>044130>05431-435>05421-425>042164>05416-420。母本的优劣顺序为：德-3>德-2、德-16>德-1>德-6、德-23>德-18>德-7>德-25>德-30>德-8>德-20>德-10。（2）单株产量特殊配合力最好的组合是：德-8×05421-425。78个组合中,产量排在前三位的组合为：德-16×044130,德-16×042164,德-23×044130（3）性状遗传力以单株角果数的广义遗传力最高,千粒重最低。试验结果结合田间表现表明：德-3、德-2和德-16这三份材料的综合性状较好,可利用其它优良材料与其杂交创造双低、高含油量、农艺性状优的材料。     

甘蓝型油菜细胞质雄性不育系105A花药败育的细胞学观察

旨在了解并揭示不育系105A的花粉败育时期和细胞学特征,为进一步正确认识细胞质雄性不育系分子机制提供必要参考依据。采用石蜡制片法,对甘蓝型油菜杂交种‘青杂5号’的雄性不育系105A及其恢复系1831R的小孢子发生和花药发育过程进行观察,以确定其花粉败育的时期和细胞学特点。研究结果表明,在不育系105A花药败育过程中,一部分发生于造孢细胞时期,属于无花粉囊败育型;另一部分发生于单核晚期,属于单核败育型,其特点为单核晚期绒毡层细胞膨大向小孢子靠近,并逐步降解,其破裂胞质退化残余物侵入药室,与小孢子混合粘连在一起,甚至有些绒毡层细胞整块脱落,成为染色很深的团块状物质,占据药室一部分空间,最终小孢子降解,花药败育。     

Production of Spontaneous Diploid Lines from Isolated Microspores Following Cryopreservation in Spring Rapeseed (Brassica napus L.)

Cryopreservation of microspores provides a potentially powerful means for increasing percentage of spontaneous diploids in microspore-derived populations in rapeseed (Brassica napus L.). This study was to investigate the production of spontaneous diploid lines in relation to steps of cryopreserving micro-spores of two spring rapeseed breeding lines G-231 and M3-124. Microspores of both genotypes were isolated and subjected to Cryopreservation. The step of cooling process (before liquid nitrogen immersion) in a Cryopreservation protocol resulted in high percentages of spontaneous diploids in lines regenerated from frozen microspores in both genotypes. Liquid nitrogen immersion and subsequent thawing process had no influence on the percentage of spontaneous diploids. A linear relationship between percentage of spontaneous diploids and cooling temperatures was detected. The lowest cooling temperature of ?40 °C yielded the highest percentages of spontaneous diploids (51.1 % in G-231 and 47.7 % in M3-124, respectively), which were more than doubled in comparison to those in the corresponding unfrozen controls. The current study indicated that cold treatment may be utilized to greatly enhance the percentage of spontaneous diploids in rapeseed breeding programs.     

Inducing male sterility in Brassica napus L. by a sulphonylurea herbicide, tribenuron-methyl

A sulphonylurea herbicide, tribenuron‐methyl, methyl 2‐[[[[(4‐methoxy‐6‐methyl‐1,3,5‐triazin‐2‐yl) methylamino] carbonyl] amino] sulphonyl] benzoate, was used to induce male sterility in rapeseed. Application of 0.2 μg tribenuron‐methyl per plant at the bolting stage with the longest floral bud <2 mm and repeated 15 days afterwards, resulted in 94.5–100% plants being male sterile in six different breeding lines, but combined with low phytotoxicity. However, excessive double application of the chemical (>0.2 μg per plant) did have some significant impact on rapeseed, including stunting, fading leaves and petals, reduction in the size of floral parts and a shortened duration of flowering. The percentage of hybrid seeds from ‘84004‘ treated with 0.2 μg × 2 tribenuron‐methyl per plant and pollinated by a male parent ‘Huaye’ was 92.7%, which met the hybridity requirement in China. The results suggest that tribenuron‐methyl could be used as an efficient chemical hybridizing agent.     

Identification of Genes Controlling Pod Length in Spring Rapeseed, Brassica napus L., and their Utilization for Yield Improvement

Experiments were undertaken to determine the inheritance of pod length in a cross with spring rapesecd, Brassica napus, and to assess the value of pod length as a criterion of selection for high seed yield. Analyses of patterns of variation in F₂; and backcross populations derived from a cross between a short-pod line TB42 and long-pod line CA553 indicated that much of the variation in pod length could be attributed to two major genes interacting in a complementary manner. Short-pods were produced when cither one or both genes were homozygous for the recessive allele. Analyses of F₃ progenies of selected F₂ and inbred-backcross lines derived from the same cross supported the two-gene hypothesis but also indicated that the effects of the major genes on pod length were possibly modified by genes of minor effect. Field testing of families derived from random intermating between F₂, plants of the TB42 × CA553 cross showed that number of pods per plant varied independently of pod length, but seed weight per pod tended to increase with increasing pod length. As a result, families with the longest pods generally had significantly higher yields than those with short pods. It was concluded that yield improvement in B. napus could be achieved through introgression of long-pod genes into cultivars with an appropriate genetic background to ensure that selection for the long-pod character would be accompanied by an increase in seed weight per pod with little or no reduction in number of pods per plant.     

砷胁迫下甘蓝型油菜苗期根、下胚轴和鲜重的全基因组关联分析

油菜是修复土壤重金属污染的理想作物,为筛选甘蓝型油菜耐砷性的显著关联单核苷酸多态性位点及相关候选基因,本研究以140份不同来源的甘蓝型油菜自交系为材料,测定和利用油菜60KSNP芯片对正常和砷胁迫条件下的相对根长(RRL)、相对下胚轴长(RHL)和相对鲜重(RFW)进行了全基因组关联分析。结果表明,与RRL、RHL和RFW显著关联的SNP位点分别为15、20和35个,单个SNP位点表型贡献率分别介于13.31%～24.39%、18.04%～33.82%和20.19%～25.06%之间;其中在A02、A07和C02染色体上同时存在与RRL、RHL和RFW显著关联的LD区间。基于油菜基因组信息在LD区间内共筛选到61个可能与砷胁迫相关的候选基因,其中PHT3;3、PHT1;9、GST、OTC5、NRAMP1和ZIP12等与重金属吸收和转运相关。实时荧光定量PCR分析结果表明, PHT3;3和PHT1;9是与甘蓝型油菜砷离子吸收转运相关的重要候选基因。本研究结果对于甘蓝型油菜耐砷胁迫机理的研究、性状的改良具有重要参考价值。     

甘蓝型油菜种皮颜色的遗传分析

应用主基因 多基因混合遗传模型并结合经典遗传学方法对甘蓝型黄籽油菜No.2127-17与黑籽油菜99Yu42和恢5148-2杂交组合的6个世代(P1,F1,P2,B1,B2和F2)群体的种子种皮颜色进行联合分析。经典遗传学表明,种皮颜色受部分显性核基因控制,并具有母体遗传特点。混合遗传分析表明,受一对加性-显性主基因 加性-显性多基因(D-1)控制,主基因对种皮颜色的表现起主要作用。其中,组合No.2127-17/99Yu42中主基因的加性和显性效应分别为7.58和2.48,而多基因则分别为1.91和1.33。在B1,B2和F2群体中种皮颜色主基因遗传率分别为35.34%,63.27%和60.76%,多基因遗传率分别为2.80%,7.12%和13.29%。环境对其遗传表现影响较大,方差占总方差的29.95%,~61.86%。     

Resynthesis of Brassica napus L. for self-incompatibility: self-incompatibility reaction, inheritance and breeding potential

Self-incompatibility (SI) in Brassica has been considered as a pollination control mechanism for commercial hybrid seed production, and so far has been extensively used in vegetable types of Brassicas. Oilseed rape Brassica napus (AACC) is naturally self-compatible in contrast to its parental species that are generally self-incompatible. Introduction of S-alleles from its parental species into oilseed rape is therefore needed to use this pollination control mechanism in commercial hybrid seed production. Self-incompatible lines of B. napus , carrying SI alleles in both A and C genomes, were resynthesized from self-incompatible B. oleracea var. italica (CC) cv.'Green Duke' and self-incompatible B. rapa ssp. oleifera (AA) cv. 'Horizon', 'Colt' and 'AC Parkland'. All resynthesized B. napus lines exhibited strong dominant SI phenotype. Reciprocal cross-compatibility was found between some of these self-incompatible lines. The inheritance of S-alleles in these resynthesized B. napus was digenic confirming that each of the parental genomes contributed one S-locus in the resynthesized B. napus lines. However, the presence of two S-loci in the two genomes was found not to be essential for imparting a strong SI phenotype. Possible use of these dominant self-incompatible resynthesized B. napus lines in hybrid breeding is discussed.     

甘蓝型油菜杂种优势和过氧化物酶的关系分析

对3个甘蓝型油菜杂交组合和亲本的农艺性状表现及其与过氧化物酶活性的关系进行了浅析,研究结果表明：杂种的产量性状,如单株粒重、单株有效角果数、有效分枝数、主花序有效角果数等,比其双亲有明显的优势;杂种花期的过氧化物酶活性高于双亲,与株高、有效分枝数、有效角果数、千粒重等性状具有显著的相关性;杂种的过氧化物酶同工酶谱表现为双亲的互补型。     

甘蓝型油菜细胞质雄性不育材料RMcms的育性变化特性的研究

广恢型细胞质雄性不育材料(RMcms)在开花后期均有不同程度的微量花粉发生，主要分布于花序的上部，在同一植株上，其微粉发生的程度次序为主花序＞一次分枝＞二次分枝；微量花粉发生与开花前18天、19天或20天的平均日最高温和平均日均温有着密切的关系，当平均日最高温稳定通过20℃、平均日均温＞14．5℃时所开花朵即有不同程度微量花粉发生，说明该类型不育材料属高温敏感型。该类型不育系的不育性因细胞质的来源不同而有很大的差异，同一不育系群体中的不同植株的不育性也存在不同程度差别。