单体附加系甜菜及空中搭载实验材料相关性状的研究

为了探明远缘杂交甜菜单体附加系M14高频传递的遗传学机理,通过胚珠分离切割、花粉败育调查和含糖量单株检测等方法,对单体附加系甜菜及空中搭载实验材料的性状进行了分析研究。研究结果验证了单体附加系甜菜M14高频传递的遗传特性存在兼性无融合生殖现象。无融合生殖体的比例达96.5%,花粉败育程度高达90%以上,大孢子母细胞通过有丝分裂形成胚囊不需要精卵结合繁育后代;有性生殖体的比例占3.5%, 有性生殖体存在致死基因的可能性。单体附加系甜菜及空中搭载的实验样品含糖量均高于普通栽培甜菜。野生白花甜菜染色体携带的高糖、抗病、无融合生殖等基因决定了远缘杂交甜菜和空中搭载实验材料具有专属的遗传学性状。研究结果为进一步开拓野生白花甜菜有益基因进行染色体工程研究奠定了良好的基础。     

单体附加系甜菜M14与异缘三倍体甜菜(VVC)核型比较分析

为了深入研究和探明单体附加系甜菜M14高频传递的机理,通过压片的方法,利用有丝分裂中期的细胞测定了染色体的绝对长度、相对长度、着丝点位置、长臂、短臂及臂比。并与异缘三倍体甜菜VVC、白花甜菜及栽培甜菜进行了染色体核型统计分析。结果表明,甜菜M14与VVC杂种细胞中附加的白花甜菜染色体绝对长度范围分别为1C：2.82±0.07 μm和2.61~3.54 μm;普通栽培甜菜绝对长度为1.94~2.89 μm,通过实验得出的最终结论是甜菜各物种的染色体是稳定的。附加的一条白花甜菜染色体上的特异基因至使单体附加系甜菜M14的高频传递特性得以表达,栽培甜菜和白花甜菜的种间杂交,通过自然重组能够将有益基因导入栽培甜菜中来。实验数据可作为分析及鉴定单体附加系甜菜M14高频传递的重要论据及其深入研究的理论基础。     

甜菜M14品系褐斑病抗性的初步研究

甜菜M14品系由于附加了一条野生白花甜菜第9号染色体,因此,研究其是否具有抗病性以及抗病程度的强弱具有十分重要的意义。通过用褐斑病原真菌感染甜菜M14品系与栽培甜菜,对比接种后叶片各种酶活性差异,并且对相关植株叶片氨基酸含量进行分析。结果表明：甜菜M14品系植株在感染褐斑病原真菌后所研究的几种酶活性都高于作为对照的栽培甜菜植株;并且甜菜M14品系植株感病后总氨基酸含量下降,而栽培甜菜感病后总氨基酸含量上升。因此,甜菜M14品系具有中度抗褐斑病的特征,更加具有研究价值。     

圆锥小麦四排穗性状基因的遗传及分子标记分析

四排穗(four-rowed spike, FRS)性状是超数小穗(supernumerary spikelets, SS)性状的一种类型,表现为在一个穗轴节片上近垂直地着生2个无柄小穗,从而增加了小穗数和穗粒数,对提高产量有一定的潜力。为了解圆锥小麦0880 FRS性状的遗传特征,将0880与正常穗(normal spike, NS)圆锥小麦0879杂交,构建了遗传群体,并对0880 (FRS) × 0879 (NS)与0879 (NS) × 0880 (FRS) F₁、F₂及F_2:3植株的穗部性状进行了调查。结果显示,正反交组合的F₁植株均表现为正常穗,F₂群体中正常穗与四排穗符合3∶1的分离比例,表明0880的四排穗性状由隐性单基因控制,将该基因定名为frs1;细胞质对frs1无显著影响。采用已定位于普通小麦A组与B组的SSR分子标记并结合混合分组分析法(BSA), 筛选出32个在双亲及F₂单株构建的四排穗型池和正常穗型池都具有多态性的SSR分子标记,利用JoinMap4.0软件构建了与frs1连锁的2A染色体11个SSR分子标记遗传图谱,其中SSR标记Xwmc598和Xwmc522位于frs1基因两侧,与该基因的遗传距离分别为4.0 cM和2.4 cM。利用2A染色体缺失系对这11个SSR进行物理定位,Xwmc598和Xwmc522均被定位在2A染色体短臂FL0.00~0.78区域。本研究的结果为frs1基因的精细定位及分子标记辅助选择奠定了基础。     

甜玉米雄性不育突变体ms2020的表型分析和基因定位

为将雄性不育基因应用于甜玉米杂交制种中，达到降低劳动成本且保证种子纯度的目的。以来源于甜玉米自交系K78的雄性不育自发突变体male sterility 2020(ms2020)为材料，构建ms2020与甜玉米自交系M08的F₁及相应的F₂遗传群体，通过表型鉴定、遗传分析和基因定位研究ms2020甜玉米雄性不育突变体。表型鉴定结果表明：F₁群体均表现为雄性可育，F₂群体出现了育性分离。不育植株能够正常抽雄，但花药不开裂、散粉异常，花药变小且颜色淡黄；1%I₂-KI染色发现不育植株的花药内包含不能正常着色的败育花粉粒。遗传分析结果表明：育性正常植株与不育植株的比例符合3∶1,表明ms2020雄性不育突变体是由单基因控制的隐性突变体。利用BSA技术，初步将目的基因定位在7号染色体短臂上；随后利用初定位区间内的20对SSR标记对不育基因进行定位，将不育基因精细定位在标记S1和W10之间，物理距离为11.30 kb。该区间内包含Zm00001d018802和Zm00001d018803...     

基于SSR分子标记的Nicotiana tobacum–N.plumbaginifolia异源染色体植株的鉴定与筛选

以烟草(Nicotiana tabacum)品种云烟87的八倍体(2n=8x=96)和野生烟草N. plumbaginifolia (2n=2x=20)的基因组DNA为模板,对340对烟草SSR引物进行筛选以获得能扩增多态性条带的引物。利用多态性引物对种间杂交后代及190株回交后代的基因组DNA进行扩增,并对N.plumbaginifolia中的SSR标记的连锁情况进行简要分析。经筛选获得了多态性引物29对。结果显示,在190株后代中, 159株的基因组DNA能扩增出N. plumbaginifolia的特异SSR位点,可以判定该159株为N. tabacum的N. plumbaginifolia异源染色体植株,其余31株植株可能不含有N. plumbaginifolia的染色体。经UPGMA聚类分析,本群体中植株的遗传多样性较为丰富,部分分子标记在后代中的出现具有完全相关性。29个标记中14个可确定来源于5条不同染色体,N.plumbaginifolia的29个位点在回交后代中的扩增效率并不相同,且效率均较低(低于31.00%),说明该杂种中N. plumbaginifolia基因组的垂直传递效率较低。利用SSR分子标记可以判定云烟87八倍体与N.plumbaginifolia杂交获得的后代为真杂种,且自该远缘杂种回交后代中筛选获得大量异源染色体植株。这些结果和筛选获得异源染色体植株为进一步创制N.tabacum-N.plumbaginifolia抗黑胫病单体附加系以及易位系奠定了基础。     

甘蓝型油菜与播娘蒿原生质体融合杂种后代的遗传研究

甘蓝型油菜(2n=38)与播娘蒿(2n=28)原生质体融合杂种F₁连续自交3代,获得F₂、F₃和F₄后代。用细胞学和SSR分子标记方法,分析杂种后代的染色体数目变异、减数分裂行为以及播娘蒿遗传成分的保留情况。结果表明在F₂、F₃和F₄代中,根尖细胞染色体平均数分别为38.47±3.17、37.65±3.23和36.66±2.95,随着自交世代增加呈减少趋势;在杂种后代减数分裂中,观察到染色体桥、染色体落后、染色体周期不同步、不均等分离等现象;杂种后代F₂、F₃和F₄代中检测到播娘蒿特征条带的平均频率分别为9.62%、2.99%和0.31%,呈减少趋势。因此要实现播娘蒿种质向油菜渗入应该重视F₂世代的选择。     

甜菜M14品系BvM14-Tpx基因克隆及原核表达体系下应答氧化胁迫功能的初步研究

为了研究甜菜M14品系BvM14-Tpx基因的抗氧化功能,本试验以带有野生白花甜菜第9号染色体的单体附加系M14品系为试验材料,利用RACE技术获得甜菜M14品系硫氧还蛋白过氧化物酶基因(BvM14-Tpx) cDNA全长,对其进行生物信息学分析,利用Real-time PCR和半定量RT-PCR技术对该基因进行组织特异性表达分析,在原核表达体系下进行BvM14-Tpx基因应答氧化胁迫研究。生物信息学分析结果表明,BvM14-Tpx基因cDNA全长为1044 bp,包含最大的ORF为489 bp,编码162个氨基酸;含有过氧化物酶Ⅱ(PrxⅡ)型的保守结构域;BvM14-Tpx蛋白与豌豆(Pisum sativum L.)和苜蓿(Medicago truncatula L.)中Tpx蛋白的亲缘性较高。组织特异性表达分析结果表明,BvM14-Tpx基因在甜菜M14品系各组织表达量从高到低的顺序是根、茎、叶、花。通过原核表达体系下BvM14-Tpx基因应答氧化胁迫的研究,表明BvM14-Tpx基因能够提高大肠杆菌对于环境中氧化胁迫的适应能力,减轻H₂O₂对细菌生长的抑制。本研究对挖掘甜菜M14品系优质基因,提高甜菜对于非生物胁迫的抗性以及开展甜菜遗传改良工作具有重要意义。     

来自野生二粒小麦的抗白粉病基因PmAS846及其染色体定位和分子标记分析

N9738是经抗性定向选择和农艺性状筛选所培育的抗白粉病普通小麦新种质,携带来自野生二粒小麦As846的抗白粉病基因PmAS846,在苗期和成株期高抗白粉菌生理小种E09和陕西关中地区流行菌系,本研究对该种质携带的抗白粉病基因进行了染色体定位和分子标记分析。对N9738和高感小麦白粉病的普通小麦品种辉县红杂交的F₁、F₂代分离群体和F_2:3代家系进行白粉病抗性鉴定和遗传分析证实,N9738苗期抗性由1个显性抗白粉病基因控制,单(缺)体分析将该基因定位在小麦5B染色体上。采用位于5B染色体的分子标记结合集群分离分析法(BSA法)分析,筛选出与PmAS846连锁的11个SSR标记和2个EST-STS标记,PmAS846两翼的SSR标记Xgwp3191和Xfcp1与该基因的遗传距离分别为7.3 cM和1.8 cM,EST-STS标记BF202652和BF482522与该基因的遗传距离均为5.1 cM。根据该基因两翼SSR标记对中国春5B染色体缺失系(Bin系)的分析将其定位在5B染色体长臂0.75~0.76区域。研究结果为PmAS846的分子标记辅助选择和精细定位奠定了基础。     

甜菜(Beta)M14花期蛋白质的变化和分析

甜菜(Beta)M14含有野生白花甜菜的染色体,这使得花期甜菜M14的蛋白质表达存有差异.本研究提取栽培甜菜和甜菜M14开花后1h的花蕾的蛋白质,利用双向电泳分离蛋白质,使用凝胶分析软件分析电泳图谱,观察到甜菜M14中有5个蛋白点存在表达差异,甜菜M14中1个蛋白点缺失,另有少量蛋白质存在表达丰度的增强或减弱.选取其中...     

2Ai-2染色体在小麦部分同源染色体代换背景中的遗传

用中间偃麦草2Ai-2染色体特异的EST-PCR标记检测5个小麦-中间偃麦草二体异代换系(包括端体代换系)与普通小麦中国春(CS)杂交后代群体,研究外源染色体2Ai-2通过杂种向后代的传递率及其结构变异,并用基因组原位杂交进行验证。结果表明,第二部分同源群不同染色体代换背景对外源染色体传递的影响不同,在2B代换系的杂种中外源染色体或片段显示优先传递,而在2D代换系的杂种中其传递力则较低,2B代换背景更有利于2Ai-2染色体或片段的传递;外源染色体在杂种后代传递过程中会发生变异,在多数组合中,变异出现在着丝粒处;与短臂相比,外源染色体长臂更容易在世代中丢失;端体代换系中的外源染色体端体在杂种后代传递过程中容易丢失,且也会发生结构变异。基因组原位杂交结果证明了分子标记跟踪外源染色体的可靠性。     

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.     

一个花粉辐射诱导的小麦-簇毛麦相互易位染色体系的分子细胞遗传学研究

利用⁶⁰Co-γ-射线处理小麦-簇毛麦6V单体添加系花粉，并给中国春授粉，在一个M1单株减数分裂中期Ⅰ检测到一个由2条小麦-簇毛麦易位染色体和一条完整小麦染色体构成的三价体，说明参与易位的2个小麦片段均来自同一条小麦染色体，推测两条易位染色体由相互易位产生。将其中涉及外源大片段的易位染色体称为外源大片段易位（large alien segment translocation, LAST），涉及外源小片段的称为外源小片段易位(small alien segment translocation, SAST)。对后代中两个易位染色体均纯合的植株（LAST’’+SAST’’, 2n = 44）进行顺次C-分带和GISH研究，结果表明外源大片段易位染色体为T7BS-6VS•6VL，外源小片段易位染色体为T6VS-7BS•7BL，易位断点分别位于7B染色体短臂约FL0.60处及6V染色体短臂约FL0.70处。在M2代群体中检测到7种染色体组成类型，比例为3(LAST’’+SAST’’)∶20(LAST’+SAST’)∶2(LAST’’+SAST’)∶1(LAST’+SAST’’)∶1LAST’∶2SAST’∶22(0型)，其中外源大片段和外源小片段易位染色体往往相伴出现。抗病鉴定结果显示抗白粉病基因位于外源大片段易位染色体T7BS-6VS•6VL上。对LAST’+SAST’型（2n = 43）M₂代单株花粉母细胞减数分裂的GISH研究结果显示，88.5%的后期Ｉ或末期Ｉ细胞中出现T6VS-7BS•7BL和T7BS-6VS•6VL的共分离。此外，在个别后期Ｉ细胞中观察到外源大片段易位染色体T7BS-6VS•6VL发生落后和着丝粒断裂现象，并在LAST’型单株（2n =42）的自交后代中筛选到一个通过着丝粒断裂-融合产生的外源小片段插入易位T7BL•6VS-7BS，这为利用外源大片段易位进一步创制携带抗病基因的小片段插入易位提供了新的思路。还分别获得了T7BS-6VS•6VL和T6VS-7BS•7BL的纯合易位系。     

Control and inheritance of resistance to yellow rust in Triticum aestivum-Lophopyrum elongatum chromosome substitution lines

A set of T. aestivum-L. elongatum chromosome substitution lines was tested for yellow rust resistance at the seedling stage. Inheritance of the resistance and esterase-5 (Est-5) variation were studied. The results demonstrated that L.elongatum carried a new gene(s) conferring yellow rust resistance. This gene was dominant and located on chromosome 3E of L. elongatum. The biochemical locus encoding Est-5was also located on chromosome 3E, and co-segregated with theYr gene(s) in the wheat background. The transmission frequencies of chromosome 3E in 3E(3A) × CS, 3E(3B) × CS and 3E(3D) × CS hybrids were scored.None of the hybrids transmitted the alien chromosome at thetheoretical maximum rate, but the transmission frequencies ofchromosome 3E in F₂ populations of 3E(3A) × CS and 3E(3D) × CS were significantly higher than in thatof 3E(3B) × CS.     

Cytological analyses of hybrids and derivatives of hybrids between durum wheat and Thinopyrum bessarabicum,using multicolour fluorescent GISH*

The wheat progenitors and other wild relatives continue to be important sources of genes for agronomically desirable traits, which can be transferred into durum wheat (Triticum turgidum; 2n = 4x = 28; AABB genomes) cultivars via hybridization. Chromosome pairing in durum × alien species hybrids provides an understanding of genomic relationships, which is useful in planning alien gene introgression strategies. Two durum cultivars, ‘Lloyd’ and ‘Langdon’, were crossed with diploid wheatgrass, Thinopyrum bessarabicum (2n = 2x = 14; JJ), to synthesize F₁ hybrids (2n = 3x = 21; ABJ) with Ph1. ‘Langdon’ disomic substitution 5D(5B) was used as a female parent to produce F₁ hybrids without Ph1, which resulted in elevation of pairing between durum and grass chromosomes – an important feature from the breeding standpoint. The F₁ hybrids were backcrossed to respective parental cultivars and BC₁ progenies were raised. ‘Langdon’ 5D(5B) substitution × Th. bessarabicum F₁ hybrids were crossed with normal ‘Langdon’ to obtain BC₁ progeny. Chromosome pairing relationships were studied in F₁ hybrids and BC₁ progenies using both conventional staining and fluorescent genomic in situ hybridization (fl‐GISH) techniques. Multicolour fl‐GISH was standardized for characterizing the nature and specificity of chromosome pairing: A–B, A–J and B–J pairing. The A–J and B–J pairing will facilitate gene introgression in durum wheat. Multicolour fl‐GISH will help in characterizing alien chromosome segments captured in the durum complement and in their location in the A and/or B genome, thereby accelerating chromosome engineering research.     

Breeding for hard red winter wheat cultivars adapted to conventional-till and no-till systems in northern latitudes

Summary Monosomic alien addition lines combining individual F. drymeja chromosomes and the L. multiflorum complement were isolated from the cross between the triploid hybrid L. multiflorum (4x) × F. drymeja (2x) and diploid L. multiflorum (2x). Chromosome pairing in the addition lines was studied at metaphase 1 of meiosis and the relationship between single F. drymeja chromosomes and the corresponding homologous pair in L. multiflorum is discussed. Trivalent frequency in the addition lines was higher than expected from observations of chromosome pairing in the triploid hybrid and there were differences between lines in the number of trivalent associations formed. There is some evidence to suggest that trivalent frequency is not entirely dependent on chromosome length and that transmission of the alien chromosome in the female is dependent on the size of the added chromosome. Morphological studies were made to assess the phenotypic effects of the addition of single F. drymeja chromosomes to the L. multiflorum complement. Two plants (2n=14) with recombination between a L. multiflorum and a F. drymeja chromosome were identified.     

Alien chromosome transmission and somatic elimination in monosomic addition lines of Gossypium australe F. Muell in G. hirsutum L.

The efficiency of using monosomic alien addition lines (MAALs) to introgress agronomical traits of interest carried by wild diploid Gossypium species into the main cultivated cotton species G. hirsutum depends on the opportunities of confronting the alien chromosome with the recipient background genome at each generation and on the occurrence of translocations and homoeologous recombinations. The selfed-progeny of five MAALs of G. australe in G. hirsutum was screened with SSR markers to determine the transmission frequency of the alien chromosome and monitor its integrity. Three MAALs revealed a transmission frequency significantly lower than the expected ratio and one MAAL presented an exclusive transmission of the additional chromosome. In these four MAAL the alien chromosome was transmitted almost unaltered. With the fifth MAAL the alien chromosome was normally transmitted but was altered in half of the plants carrying it. In one MAAL, normally carrying brown fiber, the emergence of some plants carrying white and brown fiber revealed the somatic elimination of the additional chromosome. The loss of this chromosome seems to be triggered by its deletion.