黄黑籽甘蓝型油菜类黄酮途径基因SNP位点检测分析

类黄酮物质在植物花、叶、果实和种子颜色变化的过程中起着至关重要的作用,本研究以不同黄黑籽种皮材料为研究对象,采用基因同源克隆方法,获得17个类黄酮基因全长ORF序列,在核酸和蛋白水平上分别序列差异比较表明,这些基因在不同黄黑籽材料中共存在41个不同拷贝成员。在核苷酸水平上,检测到BnTT3、BnTT18、BnTTG1和BnTTG2的单核苷酸位点数目介于16~52之间,且BnTTG2在3个不同的位置上还存在多个碱基的连续性缺失现象(119~121 bp,183~189 bp和325~330 bp),但在蛋白水平上仅存在2~16个氨基酸位点差异,说明BnTT3、BnTT18、BnTTG1和BnTTG2在不同甘蓝型黄黑籽材料中存在单核苷酸位点差异,而单核苷酸位点突变不一定导致氨基酸位点的变异。在不同黄黑籽材料中仅BnTT3和BnTT18存在一致性的氨基酸突变位点(252和87),推测BnTT3和BnTT18可能在黄黑籽甘蓝型油菜种皮颜色差异形成过程中发挥至关重要的作用。通过这些位点的等位特异PCR可以区分材料间透明种皮基因,为特异基因芯片的开发及阐明甘蓝型油菜种皮色泽性状的基因及其作用位点奠定基础。     

甘蓝型黄籽油菜的遗传研究

从1975—1990年作者及其研究集体系统坚持了甘蓝型黄籽油菜的遗传育种研究。15年来取得了以下主要研究结果: 1.甘蓝型黄籽油菜的种皮色泽不同于白菜型、芥菜型和埃塞俄比亚油菜,为土黄或姜黄,而没有纯黄,只有杂黄,即在黄色种皮上有黑色斑点、斑块或褐色环带。 2.长期自交后仍得不到遗传上稳定的纯黄后代;在大群体中,不论是自     

SRAP标记分析甘蓝型油菜多态性

采用SRAP标记对甘蓝型油菜品种1141B、垦C1、32B和32C的多态性进行了分析。每对引物组合产生13~36对比较清晰的扩增带,20对引物组合共产生419条扩增带,平均每对引物组合产生20.95条。20对引物组合共产生多态性带116条,每对引物组合产生3~10条,平均5.8条。每对引物组合产生的多态性带的比例为18.75%~38.89%,平均为28.23%。用24对引物组合对组合1141B×垦C1构建的F2群体各单株多态性进行了分析,F2群体多态性标记75条,平均为3.12条。用卡平方测验检测F2群体各单株标记基因型频率是否偏离了期望比例1∶2∶1(共显性标记)和3∶1(显性标记)的分离规律,结果表明,仅有1个SRAP标记偏离了1∶2∶1分离规律,仅占1.33%。试验结果表明,SRAP标记适于进行油菜品种的多态性分析和图谱构建,是一种经济、有效的分子标记。     

甘蓝型油菜(Brassica napus L.)千粒重性状遗传体系分析

通过遗传差异较大的2个甘蓝型油菜（Brassica napus L.）纯系亲本组合（HSTC14×宁油7号）衍生后代的世代家系群体分析，应用主基因+多基因家系世代联合分离分析方法研究油菜千粒重的遗传体系。结果表明，甘蓝型油菜HSTC14×宁油7号组合千粒重遗传体系系由一对主基因+多基因构成，主基因中只有加性效应(d = 0.1062),不存在显     

RAPDs和RFLPs分析甘蓝型杂交油菜亲本的遗传多样性

利用RAPD和RFLP分子标记技术对甘蓝型杂交油菜亲本遗传多样性的分析结果表明:(1)20个亲本间具有丰富的遗传多样性,40个引物扩增出277条多态性带,其中21条带为10个亲本所特有,12个探针得到了117条多态性杂交带,其中7条带为5个亲本所特有;(2)不育系与恢复系之间的遗传差异大于不育系内和恢复系内的遗传差异;(3)依RAPDs与RFLPs估     

白芥和甘蓝型油菜属间杂种后代种子结构比较

白芥具有很多优良的农艺性状,从白芥和甘蓝型油菜属间体细胞杂种后代中筛选出多个具有黄籽或趋向黄籽性状的株系,利用光学显微镜和电子显微镜技术观察它们种子的结构。回交后代种皮解剖结构与甘蓝型油菜相似,而与白芥相差较远。种皮色素主要分布在栅栏层,甘蓝型油菜和部分后代株系中有色素分布,而白芥和部分黄籽后代株系中没有色素分布。栅栏层在甘蓝型油菜中最厚,在白芥中最薄,而后代介于两者之间。回交后代和甘蓝型油菜种皮表面纹饰为网-穴状,白芥为沟槽状或水疱状。胚子叶细胞面积以白芥最小,甘蓝型油菜最大,后代介于两者之间;而蛋白体面积指数以白芥最大,甘蓝型油菜最小,后代介于两者之间。超微结构观察表明,亲本和后代蛋白体均为球状晶体蛋白体,油体有大、小两种,其大小在亲本和后代间有差异。上述结果表明,回交后代株系种子解剖结构与甘蓝型油菜相近,种皮颜色、色泽深浅和栅栏层厚度,以及胚子叶细胞大小、油体和蛋白体等受亲本白芥的影响而发生变化。     

甘蓝型油菜种皮花青素相对含量测定方法研究

在配合甘蓝型油菜育种工作中,花青素含量的变化是重要的一个测定指标,笔者介绍了一种改进的测定甘蓝型油菜种皮中花青素相对含量的方法pH差示法,同常用的盐酸甲醇法相比较,同样不需要标准样品,成本低廉,测定简便快捷,同时能有效的去除甘蓝型油菜种皮中褐色物质对测定结果的干扰,所得结果能更准确地表现出不同色泽的种皮中花青素含量的变化。pH差示法在简便,快捷的基础上,所测定甘蓝型油菜种皮花青素相对含量更为准确、有效。     

SSR和SRAP标记揭示甘蓝型油菜遗传多样性的差异分析

分别用SSR和SRAP标记分析了来自国内外的19个甘蓝型油菜（Brassicanapus L.）品种间的遗传距离及其与选育年代的相关关系,并构建聚类图。从726个SRAP引物组合中筛选出23对多态性较好的组合扩增得到了234个多态性标记;35对SSR引物获得了138条多态性标记。结果显示,SSR标记揭示的遗传距离大于SRAP标记。SSR揭示的国内甘蓝型油菜品种间平均遗传距离大于国外品种,而SRAP标记得到的结果与SSR标记分析所得结果相反。SSR平均遗传距离与育成年代的相关性较小,SRAP平均遗传距离与育成年代存在一定的负相关,其相关系数为-0.34。研究认为,SRAP标记更适用于通过分析遗传距离以获得杂种优势;从种质资源保存的角度出发,使用SSR标记能较全面保证种质资源遗传多样性;结合SSR标记和SRAP标记即能从全基因组又能有重点的从功能基因组分析育种材料的遗传多样性,对于中长期的育种目标有较大的意义。     

cDNA-SRAP标记在玉米氮素营养诊断上的应用初探

为探讨对玉米氮素营养状况进行分子诊断的可行性,以大面积推广的玉米( Zea mays)杂交种郑单958为材料,用cDNA-SRAP标记的方法研究了玉米幼苗在不同氮素水平下基因表达的差异,对其中的一些差异表达片段进行了克隆和测序,并通过Real-time PCR鉴定了克隆的差异表达片段的真实性.结果表明:不同的SRAP标...     

芥菜型油菜黄黑种皮多酚差异的紫外-可见光谱研究

以芥菜型自交系黄黑油菜种皮为材料,研究了其多酚提取液的紫外-可见吸收光谱, 测定了芥菜型油菜黄黑种皮样以没食子酸和茶儿茶素标准计的多酚含量,发现黄种皮的多酚含量约为黑种皮的90%,显著区别于甘蓝型油菜的10%,因此多酚含量高可能是芥菜型黄籽油菜性状稳定的根本内因。基于香草醛试验,推测黑种皮中含有3-OH的黄烷醇类多酚,而黄种皮中没有。基于加入位移试剂后特征吸收谱带Ⅰ与Ⅱ的位移方向与大小,推测黄酮醇类多酚A环在黄种皮中可能主要为单酚羟基,而在黑种皮中可能主要为邻二酚羟基结构。这种多酚分子的结构差异可能是形成芥菜型油菜黄黑种皮色泽差异的主要原因。     

甘蓝型黄籽油菜性状组间的典型相关分析

以7个不同遗传来源的甘蓝型黄籽品系所配制的完全双列杂交作样本,对甘蓝型黄籽油菜进行典型相关分析。结果表明,在性状组间关联上起主要作用的是株型性状中的株高、产量性状中的单株粒重、品质性状中的皮壳率和黄籽度。用多目标综合选择指数,讨论了甘蓝型黄籽油菜的选择问题。     

Development of yellow-seeded Brassica napus of double low quality

Two yellow‐seeded white‐petalled Brassica napus F₇ inbred lines, developed from interspecific crosses, containing 26–28% emcic acid and more than 40 μmol glucosinolates (GLS)/g seed were crossed with two black/dark brown seeded B. napus varieties of double low quality and 287 doubled haploid (DH) lines were produced. The segregation in the DH lines indicated that three to four gene loci are involved in the determination of seed colour, and yellow seeds are formed when all alleles in all loci are in the homozygous recessive state. A dominant gene governed white petal colour and is linked with an erucic acid allele that, in the homozygous condition, produces 26–28% erucic acid. Four gene loci are involved in the control of total GLS content where low GLS was due to the presence of recessive alleles in the homozygous condition in all loci. From the DH breeding population a yellow‐seeded, yellow‐petalled, zero erucic acid line was obtained. This line was further crossed with conventional B. napus varieties of double low quality and, following pedigree selection, a yellow seeded B. napus of double low quality was obtained. The yellow seeds had higher oil plus protein content and lower fibre content than black seeds. A reduction of the concentration of chromogenic substances was found in the transparent seed coat of the yellow‐seeded B. napus.     

Maternal and embryonal control of seed colour by different Brassica alboglabra chromosomes

Most oilseed rape, Brassica napus, cultivars are black‐seeded. The progenitor species, Brassica rapa, has either yellow or black seeds, while known cultivars of the other progenitor species Brassica oleracea/alboglabra have black seeds. To determine which chromosomes of B. alboglabra are carriers of seed colour genes, B. rapaalboglabra monosomic addition lines were produced from a B. napus resynthesized from yellow‐seeded B. rapa and brown/black‐seeded B. alboglabra. Eight out of nine possible lines have been developed and transmission frequencies of the alien chromosomes were estimated. Three B. alboglabra chromosomes in three of these lines influenced seed colour. B. rapa plants carrying alien chromosome 1 exhibited a maternal control of seed colour and produced only brown seeds, which gave rise to plants with either yellow or brown seeds. However, B. rapa plants carrying alien chromosome 4 or another as yet unidentified alien chromosome exhibited an embryonal control of seed colour and produced a mixture of yellow and brown seeds. The yellow seeds gave rise to yellow‐seeded plants, while the brown seeds gave rise to plants that yielded a mixture of yellow and brown seeds, depending on the absence or presence, respectively, of the B. alboglabra chromosome. Consequently, both maternal and embryonal control of seed colour are expected to contribute to the black‐seeded phenotype of oilseed rape.     

The production of yellow-seeded Brassica napus (AACC) through crossing interspecific hybrids of B. campestris (AA) and B. carinata (BBCC) with B. napus

To transfer the genes for yellow seed coat from both genomes A and C to B. napus (AACC), the hexaploid of Brassica (AABBCC) was synthesised from reciprocal interspecific crosses between yellow-seeded B.campestris (AA) and B.carinata (BBCC). The hexaploid with 27 pairs of chromosomes was red-seeded which showed that genic interaction existed in the trigenomic plants for the colour of the seed coat. Hundreds of hybrid seeds were obtained from crosses between the red-seeded hexaploid and partial yellow or brown-seeded varieties of B. napus as pollen donor. The majority of the hybrid plants (AABCC) were self fertile with brown seeds. It appeared that the chromosomes of the B genome were excluded during the meiosis of the pentaploid and a high proportion of the genetically balanced AC gametes could be produced. The fertility of the F2 population was increased and even reached normal levels for some plants. Seventy-three plants with the yellow-seeded character were isolated from 2590 open-pollinated F2 plants, most with increased fertility. After two successive self-pollinations, 18 lines produced yellow seeds and no brown seeds segregated from these populations. The morphology of the novel yellow-seeded plants was basically towards B. napus. Esterase isoenzyme electrophoresis showed that the plants contained some of the genetic background of B. campestris, B. carinata and B. napus. Cytological analysis has shown that at least some yellow-seeded lines have the B.napus AACC genome composition with 38 chromosomes and normal meiotic pairing. This revised version was published online in July 2006 with corrections to the Cover Date.     

甘蓝型油菜种皮黑色素形成机理的研究

甘蓝型油菜在种子发育过程中种皮中黑色素含量与多酚和游离酪氨酸呈极显著或显著负相关. 提纯后黑色素的HPLC分析结果表明黑籽和黄籽油菜种皮黑色素的组成相同. 在种子发育后期, 种皮黑色素的增加速率(0.255)与游离酪氨酸(0.071)和多酚(0.208)含量的下降速率之和(0.279)相近, 说明黑籽油菜中黑色素是以游离酪氨酸和多酚为前体     

甘蓝型油菜成熟籽粒DNA快速提取方法探讨

以室温保存不同年份的甘蓝型油菜种子为材料,采用改良的CTAB法和DNA抽提试剂盒分别提取甘蓝型油菜籽粒DNA,探讨适合甘蓝型油菜籽粒DNA提取的方法。结果表明,不同保存年份种子提取的DNA含量无显著差异;受种皮色泽的影响也较小;用改良的CTAB法比试剂盒所提DNA产量高,结构完整性好,成本较低。PCR扩增检测结果表明两种方法获得的DNA均能满足一般的分子实验。因此,从油菜干籽粒中直接提取DNA可以节约时间,显著提高工作效率,为快速高效地进行甘蓝型油菜种质资源和遗传多样性分析奠定了基础,同时,也利于快速鉴定推广品种是否带有外源基因。     

118份春性甘蓝型油菜恢复系的桔红花色位点鉴定

甘蓝型油菜桔红花色恢复系在杂交制种过程中起到指示作用,可去除杂株,因此培育出桔红花色优良恢复系是十分有必要的。本研究利用已开发的与桔红花色位点Bnpc1和Bnpc2紧密连锁的4个分子标记对118份春性甘蓝型油菜恢复系的桔红花色位点基因型进行鉴定,从中筛选出Bnpc1位点为显性,Bnpc2位点为隐性(BnPC1 BnPC1 Bnpc2 Bnpc2)或Bnpc1位点为隐性,Bnpc2位点为显性(Bnpc1 Bnpc1 BnPC2 BnPC2)的纯合黄花资源,从而可以利用这两种基因型恢复系相互杂交,选育出桔红花色优良恢复系。这为培育出桔红花色优良恢复系提供了一种简单有效的方法,同时也为花色选育提供了优良桔红花色遗传资源。     

Production of yellow-seeded Brassica napus through interspecific crosses

Yellow‐seeded Brassica napus was developed from interspecific crosses between yellow‐seeded Brassica rapa var.‘yellow sarson’ (AA), black‐seeded Brassica alboglabra (CC), yellow‐seeded Brassica carinata (Bbcc) and black‐seeded B. napus (AACC). Three different interspecific crossing approaches were undertaken. Approaches 1 and 2 were designed directly to develop yellow‐seeded B. napus while approach 3 was designed to produce a yellow‐seeded CC genome species. Approaches 1 and 2 differed in the steps taken after trigenomic interspecific hybrids (ABC) were generated from B. carinata×B. rapa crosses. The aim of approach 1 was to transfer the yellow seed colour genes from the A to the C genome as an intermediate step in developing yellow‐seeded B. napus. For this purpose, the ABC hybrids were crossed with black‐seeded B. napus and the three‐way interspecific hybrids were self‐pollinated for a number of generations. The F₇ generation resulted in the yellowish‐brown‐seeded B. napus line, No. 06. Crossing this line with the B. napus line No. 01, resynthesized from a black‐seeded B. alboglabra x B. rapa var.‘yellow sarson’ cross (containing the yellow seed colour genes in its AA genome), yielded yellow‐seeded B. napus. This result indicated that the yellow seed colour genes were transferred from the A to the C genome in the yellowish‐brown seed colour line No. 06. In approach 2, trigenomic diploids (AABBCC) were generated from the above‐mentioned trigenomic haploids (ABC). The seed colour of the trigenomic diploid was brown, in contrast to the yellow seed colour of the parental species. Trigenomic diploids were crossed with the resynthesized B. napus line No. 01 to eliminate the B genome chromosomes, and to develop yellow‐seeded B. napus with the AA genome of ‘yellow sarson’ and the CC genome of B. carinata with yellow seed colour genes. This interspecific cross failed to generate any yellow‐seeded B. napus. Approach 3 was to develop yellow‐seeded CC genome species from B. alboglabra×B. carinata crosses. It was possible to obtain a yellowish‐brown seeded B. alboglabra, but crossing this B. alboglabra with B. rapa var.‘yellow sarson’ failed to produce yellow seed in the resynthesized B. napus. The results of approaches 2 and 3 demonstrated that yellow‐seeded B. napus cannot be developed by combining the yellow seed colour genes of the CC genome of yellow‐seeded B. carinata and the AA genome of ‘yellow sarson’.