一种鉴定油菜远缘杂种的细胞学方法

在众多的染色体观察技术中,压片法由于其步骤简单、操作方便等特点,在油菜及其近线种属植物染色体的研究中,得到了广泛应用（蓝泽蓬等,1994）。然而,通常观察的部位为种子根尖,由于远缘杂交给实率普遍较低,得到远缘杂种又极其珍贵,取其种子根尖来鉴定是否为真杂种则相当困难,同时,在鉴定附加系、单体与缺体等非整信体时,取根尖观察也存在相似问题。如通过组培技术,来增加观察部位的数量,则在技术上要求过高,不是一种最理想的方法。我们采用幼小雌蕊观察法,能快速地鉴定当代单株的染色体组成。本文对以上三种方法进行了比较分…     

用RAPD和染色体原位杂交检测Elymus rectisetus染色体附加到普通小麦中

报道小麦×Elymusrectisetus属间杂种BC2F236个衍生系,通过染色体原位杂交和RAPD分析,检测Elymusrectisetus染色体及其特异染色体DNA标记。原位杂交结果表明,36个衍生系中大多数系含1对异源Erectisetus染色体,少数系含3条以上Ereclisetus染色体;RAPD结果表明,有13个随机引物（OPBA-08、OPB－14OPEA-09、OPF-05、OPF－09、OPJ-05、OPK-03、OPN-12、OPP-20、OPS－12、OPT-20、OPZ-09、OPZ-11）能够在这36个衍生系中分别扩增出普通小麦所没有的Erectisetus特异的染色体DNA片段。其中Erectisetus特异染色体DNA片段OPF－05480bp、OPF－09650bp和OPZ-11350bp只能够在“1048”系统的全部8个株系中扩增出;OPN-12350bp只能够在“1057”系统的全部6个株系中扩增出;OPB-14900bp和OPM-05420bp只能够在“1034”、“1026”2个系统的全部22个株系和“1057”－5－3株系中扩增出。直接获得了3个类型不同的异附加系,省去常规通过测配和附加系两两杂交F1PMCMI细胞学鉴定来确证异附加系之间的异同。由此说明染色体原位杂交和RAPD分析有机结合是鉴定异附加系（异代换系）快速有效的方法,且方法简单、易行．     

应用RAPD标记法鉴定甘蓝型油菜杂种纯度

近年 ,华中农大作物遗传改良国家重点实验室国家油菜改良武汉分中心的在职研究生段院和刘平武等 ,对用RAPD标记鉴定甘蓝型油菜杂种H990 9的纯度作了研究和报道。他们对杂交组合H990 9两亲本的指纹图谱作了分析研究 ,选出S374、SI334、SI36 5、UBC5 2 34个引物作杂种的纯度鉴定     

一个小麦—黑麦染色体代换的细胞学鉴定

对从六倍体小黑麦与普通小麦的杂种后代中获得的矮秆抗病选系84056-1-36-1进行体细胞C-分带鉴定,结果表明,它的21对染色体中,有1对短臂带型与1R相似的黑麦染色体代换了小麦的1D。观察“中国春”双端二体1A、1B与该选系杂种F_1的花粉母细胞染色体配对,发现分别有82.56%和65.71%的细胞出现异型三价体,所有细胞至少有两个形态不同的单价体;而在“中国春”端二体IDL与84056-1-36-1的杂种中,端体不配对的花粉母细胞占100%,经C-分带后,另外1条单价体显示明显的端带。从上述这些结果推断84056-1-36-1为1R(1D)代换系。     

芥菜型油菜与埃塞俄比亚芥杂种基因组原位杂交分析

原产于非洲的埃塞俄比亚芥(Brassica carinata,2n=34,BBCC)具有适应于炎热干旱地区种植等特点,是改良我国芥菜型油菜(B.juncea,2n=36,AABB)的重要种质资源。本研究用基因组原位杂交方法(GISH,Genomic in situ hybridization)分析了芥菜型油菜与埃塞俄比亚芥种间杂种花粉母细胞的染色体分离,发现在后期I染色体主要以17∶18类型分离,其次是16∶19,染色体落后现象偶然发生,其中B染色体组以8∶8的分离比率较高,表明不同来源的B染色体可正常配对分离。本研究表明,芥菜型油菜与埃塞俄比亚芥远缘杂交,通过染色体同源重组(B染色体间),以及部分同源染色体配对交换的方式(A、B、C基因组间),有可能将埃塞俄比亚芥优良遗传成分转移到芥菜型油菜中。     

RAPD分析─鉴定柑桔体细胞杂种的快速方法

本文利用改进的DNA提取方法,从Volkamer柠檬(Citrus volkameriana Ten. and Pasq.)和酸橙(C. aurantium L.)及其原生质体杂种植株的叶片中抽提总DNA,进行RAPD(Random Amplified Polymorphic DNA)分析。结果表明: 在随机选取的15种引物中,有10种可单独或与其它引物一道鉴定这一组合的体细胞杂种。与形态学性状观察、同工酶及ONA杂交分析等方法比较,RAPD分析是一种可在试管苗期即可直接、准确、快速鉴定柑桔体细胞杂种的方法。     

抗白粉病小麦——中间偃麦草异附加系的细胞学和RAPD鉴定

利用细胞学和ＲＡＰＤ技术,对从小麦与中间偃麦草杂种后代中选育的抗白粉病异附加系ＤＡＬ６６进行了鉴定。结果证明ＤＡＬ６６根尖细胞染色体数为４４,花粉母细胞减数第一分裂中期（ＰＭＣ ＭＩ）杂色体模型为２ｎ＝２２Ⅱ。对ＤＡＬ６６及其双亲进行ＲＡＰＤ分析,从４０个随机引物中筛选出１个特异引物（ＯＰＥ－０２）能够稳定地扩增出特异带型。     

远缘花粉延迟授粉诱导小麦单性生殖的染色体观察

应用很尖细胞染色体制片技术,观察了以普通小麦品种间杂种第一代为母本,以硬粒小麦（T,durum)品种“阿普洛”等和黑麦（Secale cereale）品种“D. T. R.”等的花粉为诱导者授粉产生的后代染忿2术－从中筛选鉴定出真正为单性生殖的后代,可作为选育品种的材料。     

大麦×小麦杂种愈伤组织及再生植株的细胞学观察

本文研究了Ant-13大麦×中国春小麦未成熟胚诱导的愈伤组织细胞,再生植株体细胞和花粉母细胞染色体的变化情况。发现在这三个不同分化和发育阶段中都存在着混倍体现象。但随着分化和发育过程的进行,混倍体程度越来越小,正常双单倍体细胞的比例越来越大。从再生植株花粉母细胞减数分裂前的有丝分裂开始到整个减数分裂过程中都可以看到染色体行为的异常现象,从而形成败育花粉,造成杂种不孕。花粉败育发生在单核小孢子时期。     

小麦和无融合生殖披碱草杂交后代(BC2F2)的无融合生殖及胚胎发育过程中的异常现象研究

对小麦（Triticum aestivum）和无融合生殖披碱草（Elymus rectisetus）的染色体数目为42的杂种后代（BC2F2）单株进行了RAPD检测和胚胎学研究,RAPD检测结果表明：染色体数目为42条的BC2F2单株的遗传组成与普通小麦的遗传组成十分接近,但是在部分单株中出现了披碱草的特异带。由此可以推测,经过回交和自交后小麦草的部分染色体片段已经整合进了小麦的染色体。在部分BC2F2单株胚胎学切片中发现了较高比例的（5%左右）双孢原、早发胚以及多胚囊等无融合生殖现象,直接表明了无融合生殖基因转移。由于基因整合的多样性。无融合生殖基因在有些单株中并没有充分表达,从而造成了某些单株胚胎发育的异常。     

拟三属间杂交品系神农高糖蔗种质来源鉴定

新品种选育的关键在于优异种质资源的创制和利用,远缘杂交是创新种质的重要手段。近年来,有机构宣称培育了一种由高粱、玉米、甘蔗属间杂交而成的神农高糖蔗新品系。为了准确地鉴定其种质来源,本研究利用形态学、SCo T标记和GISH技术对神农高糖蔗进行了研究。形态学初步鉴定结果表明:从根、茎、叶等不同器官的形态特征来看,神农高糖蔗与甘蔗、玉米、高粱都具有较多相似之处,无法准确地甄别出神农高糖蔗是否为真杂种。根据SCo T标记的聚类结果,可以将12份供试材料分成I、II、III 3大类,I类包括3份高粱材料和神农高糖蔗,6份甘蔗材料均聚为II类,2份玉米材料聚为III类。在整个聚类图中神农高糖蔗和非洲甜高粱的相似系数最大,说明神农高糖蔗与非洲甜高粱的遗传距离较小,它们之间存在较小的遗传差异。比较基因组原位杂交结果表明,神农高糖蔗只含有甜高粱的血缘,而不含有甘蔗和玉米的血缘。本研究结合3种鉴定方法,可以基本确定神农高糖蔗并非甘蔗、高粱和玉米的属间杂种,而是甜高粱的一个品系(品种)。本研究结果不仅揭露了虚假报道,避免不必要的损失,还探讨了甜高粱作为甘蔗远缘杂交种质资源的利用前景。     

Determination of the frequency of wheat-rye chromosome pairing in wheat x rye hybrids with and without chromosome 5B

Genomic in-situ hybridization (GISH) was used to determine the amount of wheat-rye chromosome pairing in wheat (Triticum aestivum) x rye (Secale cereale) hybrids having chromosome 5B present, absent, or replaced by an extra dose of chromosome 5D. The levels of overall chromosome pairing were similar to those reported earlier but the levels of wheat-rye pairing were higher than earlier determinations using C-banding. Significant differences in chromosome pairing were found between the three genotypes studied. Both of the chromosome-5B-deficient hybrid genotypes showed much higher pairing than the euploid wheat hybrid. However, the 5B-deficient hybrid carrying an extra chromosome 5D had significantly less wheat-rye pairing than the simple 5B-deficient genotype, indicating the presence of a suppressing factor on chromosome 5D. Non-homologous/non-homoeologous chromosome pairing was observed in all three hybrid genotypes. The value of GISH for assessing the level of wheat-alien chromosome pairing in wheat/alien hybrids and the effectiveness of wheat genotypes that affect homoeologous chromosome pairing is demonstrated.     

一个多抗小麦—中间偃麦草新种质的选育和细胞分子生物学鉴定

经过多年田间和温室接种抗病性鉴定,从(77-5433×中5)杂交组合花药培养后代中选育出一个兼抗大麦黄矮病、条锈、叶锈和秆锈4种小麦主要病害的新种质遗4212。遗4212的体细胞染色体数为42,在减数分裂中期Ⅰ,在几乎所有的花粉母细胞中都可以观察到21个二价体,这说明遗4212是一个在遗传上业已稳定的整倍体材料。对(遗4212×77-5433)F_1代花粉母细胞的观察表明,遗4212可能是含1对外源中间偃麦草染色体的代换系或具较大中间偃麦草染色体片段的易位系。用基因组原位杂交(genomic in situ hybridization,GISH)对遗4212的有丝分裂中期相、减数分裂后期Ⅰ相和(遗4212×77-5433)F_1代花粉母细胞减数分裂中期Ⅰ、后期Ⅰ进行了检测,确证遗4212含1对外源中间偃麦草染色体。这些结果表明,遗4212是一个小麦一中间偃麦草代换系,其抗病性来自其携带的1对中间偃麦草。     

玉米和类玉米种间杂交后代细胞学鉴定

利用组成玉米异染色质钮的180-bp重复序列和TR-1元件以及45S rDNA对玉米自交系F107、GB57、二倍体多年生类玉米及其远缘杂交后代的染色体进行荧光原位杂交,确定了3种重复序列在亲本染色体上的分布;同时对远缘杂交后代进行了细胞学鉴定,通过荧光信号在染色体上的位置,证实远缘杂交后代中异源种质的染色体来源;讨论了异染色质钮重复序列对玉米和其野生种杂交后代外源染色体整合和染色体行为等方面研究的应用。     

基因组原位杂交的新进展及其在植物中的应用

基因组原位杂交 ( Genomic in situ hybridization GISH)是 2 0世纪 80年代末发展起来的一种原位杂交技术。它最初应用于动物方面的研究[1 ] ,但很快被植物方面所借用 ,并且使用频率高于动物方面的研究。它采用来自一个物种的总基因组 DNA作为标记探针 ,用另一物种的总基因组 DNA以适当的浓度进行封阻 ,在靶染色体上进行原位杂交。在封阻DNA和标记 DNA探针之间 ,封阻 DNA优先与一般序列杂交 ,剩下的特异性序列主要被标记探针所杂交。在此基础上 ,人们先后发展了荧光基因组原位杂交、多色基因组原位杂交和比较基因组原位杂交等技术 ,…     

Cytological and molecular characterization of oat x maize partial hybrids

In cereals, interspecific and intergeneric hybridizations (wide crosses) which yield karyotypically stable hybrid plants have been used as starting points to widen the genetic base of a crop and to construct stocks for genetic analysis. Also, uniparental genome elimination in karyotypically unstable hybrids has been utilized for cereal haploid production. We have crossed hexaploid oat (2n=6x=42, Avena sativa L.) and maize (2n=2x=20, Zea mays L.) and recovered 90 progenies through embryo rescue. Fifty-two plants (58%) produced from oatxmaize hybridization were oat haploids (2n=3x=21) following maize chromosome elimination. Twenty-eight plants (31%) were found to be stable partial hybrids with 1–4 maize chromosomes in addition to a haploid set of 21 oat chromosomes (2n=21+1 to 2n=21+4). Ten of the ninety plants produced were found to be apparent chromosomal chimeras, where some tissues in a given plant contained maize chromosomes while other tissues did not, or else different tissues contained a different number of maize chromosomes. DNA restriction fragment length polymorphisms (RFLPs) were used to identify the maize chromosome(s) present in the various oat-maize progenies. Maize chromosomes 2, 3, 4, 5, 6, 7, 8, and 9 were detected in partial hybrids and chromosomal chimeras. Maize chromosomes 1 and 10 were not detected in the plants analyzed to-date. Furthermore, partial self-fertility, which is common in oat haploids, was also observed in some oat-maize hybrids. Upon selfing, partial hybrids with one or two maize chromosomes showed nearly complete transmission of the maize chromosome to give self-fertile maize-chromosome-addition oat plants. Fertile lines were recovered that contained an added maize chromosome or chromosome pair representing six of the ten maize chromosomes. Four independently derived disomic maize chromosome addition lines contained chromosome 4, one line carried chromosome 7, two lines had chromosome 9, one had chromosome 2, and one had chromosome 3. One maize chromosome-8 monosomic addition line was also identified. We also identified a double disomic addition line containing both maize chromosomes 4 and 7. This constitutes the first report of the production of karyotypically stable partial hybrids involving highly unrelated species from two subfamilies of the Gramineae (Pooideae — oat, and Panicoideae — maize) and the subsequent recovery of fertile oat-maize chromosome addition lines. These represent novel material for gene/ marker mapping, maize chromosome manipulation, the study of maize gene expression in oat, and the transfer of maize DNA, genes, or active transposons to oat.Joint contribution of the Minnesota Agricultural Experiment Station and USDA-ARS. Scientific journal series paper No. 21 859 of the Minnesota Agricultural Experiment Station. Mention of a trademark or proprietary product does not constitute a guarantee or warranty by the USDA-ARS or the University of Minnesota and does not imply approval over other products that also may be suitable     

Visualization of Secale cereale DNA in wheat germ plasm by fluorescent in situ hybridization

Homozygous wheat/rye (1BL/1RS or 1AS/ 1RL) translocation lines have significantly contributed to wheat production, and several other wheat/rye translocation lines show a potential promise against biotic and abiotic stresses. Detecting the presence of rye at the chromosome level is feasible by C-banding and isozyme protocols, but the diagnostic strength of genomic in situ hybridization for eventually analyzing smaller DNA introgressions has greater significance. As a first step we have applied the genomic in situ hybridization technique to detect rye chromosomes in a wheat background using germ plasm of agricultural significance. By this method rye contributions to the translocations 1BL/1RS, 1AL/1RS, 5AS/5RL and 6BS/6RL could be identified. Differential labelling has further enabled the detection of rye and Thinopyrum bessarabicum chromosomes in a trigeneric hybrid of Triticum aestivum/Th. bessarabicum//Secale cereale.