小麦-中间偃麦草2A/6St代换系014-459的分子细胞遗传学鉴定

中间偃麦草是小麦遗传改良的有用资源,育成了大量的小麦-中间偃麦草附加系、代换系及部分双二倍体。中4是小麦-中间偃麦草部分双二倍体,很方便与普通小麦杂交并被广泛用于小麦的遗传改良。014-459是中4与普通小麦杂交后代,具有一些特殊特性,材料014-459与一些普通小麦杂交,无论正反交,其F1表现为不育,而与另一些普通小麦的杂交F1表现为可育,此外,材料014-459粗蛋白和湿面筋含量很高。基于014-459的这些特性,猜测其可能具有中间偃麦草染色体片段,为此对014-459进行了细胞学鉴定。FISH和GISH以及PLUG标记分析用来分析材料014-459的染色体组成情况。连续的FISH和GISH试验证实小麦-中间偃麦草部分双二部体中4与小麦杂交后代品系014-459的1对小麦2A染色体被来自中间偃麦草的1对St染色体代换, PLUG标记分析证实这1对St染色体属于第6同源群,可能来自中间偃麦草的St染色体被代换进小麦中造成了材料014-459的一些特性。对品系014-459的分子细胞遗传学鉴定对促进中间偃麦草6St染色体在小麦中利用及小麦品质改良有积极作用。     

Fluorescent in situ hybridization — a useful aid to the introduction of alien genetic variation into wheat

Summary Fluorescent in situ hybridization (FISH) of DNA to plant chromosomes has proved to be a powerful cytogenetic tool. The value of fluorescent in situ hybridization of total genomic DNA (GISH) of related species is demonstrated in the determination of wheat/alien chromosome pairing in hybrids. Its use for assessing the relative merits of the various genes that affect chromosome pairing is also shown.The ability of GISH to identify the presence in wheat of whole alien chromosomes or alien chromosome segments is illustrated. The potential of FISH for detecting repeated DNA sequences, low copy sequences and single copy genes is discussed.Abbreviations FISH fluorescent in situ hybridization - GISH genomic in situ hybridization - PRINS primer-induced in situ hybridization     

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.     

对基因组原位杂交信号释译可能出现的片面性——来自一个小麦易位系(A-3)中外源遗传物质鉴定的启示

利用基因组原位杂交(GISH)和种子醇溶蛋白电泳(A-PAGE)技术对一个可能携带有10倍体长穗偃麦草(Thinopyrum ponticum (Host) Liu & Wang)遗传物质的小麦新种质A-3进行了综合鉴定. 用普通小麦"中国春"(Triticum aesticum L. cv. Chinese Spring)基因组(ABD)DNA作探针, 拟鹅观草[Pseudoroegneria stipifolia(Czern e     

Molecular cytogenetic characterization of Thinopyrum genomes conferring perennial growth habit in wheat- Thinopyrum amphiploids

Seven wheat‐Thinopyrum amphiploids, AT 3425, AgCs, PI 550710, PI 550711, PI 550712, PI 550713 and PI 550714, were evaluated for perennial growth habit in the field. Three of them, AgCs, AT 3425, and PI 550713, were identified as perennials. Fluorescent genomic in situ hybridization (FGISH) patterns of mitotic chromosomes indicated that AgCs had seven pairs of Thinopyrum chromosomes and 21 pairs of wheat chromosomes. PI 550713 and AT 3425 showed similar FGISH patterns of mitotic chromosomes with three pairs of wheat‐Thinopyrum translocated chromosomes, seven pairs of Thinopyrum chromosomes, and 18 pairs of wheat chromosomes. Thinopyrum chromosome pairing in the Fi hybrid of AT 3425 with AgCs demonstrated differences between Thinopyrum genomes in these two amphiploids. Based on chromosome constitutions, pairing and reported pedigrees, AgCs and AT 3425 were identified as a wheat‐Thinopyrum elongatum amphiploid and partial wheat‐Thinopyrum ponticum amphiploid, respectively. Chromosome pairing in the F₁ hybrid between AT 3425 and PI 550713 revealed that these two amphiploids contained the same Thinopyrum genome. Two different Thinopyrum genomes conferring perennial growth habit were identified from the perennial amphiploids and characterized cytogenetically.     

十个八倍体小偃麦的细胞学鉴定和染色体构成分析

培育新的八倍体小偃麦,对于利用偃麦草遗传物质进行小麦的遗传改良具有重要意义。本研究利用细胞学和基因组原位杂交技术,对从中间偃麦草与小麦品种烟农15杂交后代选育出的10个八倍体小偃麦山农TE256、山农TE259、山农TE261、山农TE262、山农TE263、山农TE265、山农TE266、山农TE267-1、山农TE270和山农TE274进行了细胞学鉴定和染色体构成分析。结果表明,10个八倍体小偃麦绝大多数单株根尖细胞的染色体数目为2n=56,个别单株含有54或55条染色体;大多数2n=56单株的花粉母细胞在减数分裂中期I的染色体构型为2n=28II,少数花粉母细胞存在单价体、三价体或四价体,后期I染色体可均等分向两极,仅有极少数细胞出现染色单体提前分离等现象;10个八倍体小偃麦均含有普通小麦的全套染色体和中间偃麦草的1个混合染色体基组,其中间偃麦草染色体是由来自中间偃麦草3个不同染色体基组的染色体构成的混合染色体基组,其染色体构成分别为2St+8J~S+2J+2J-St、2St+8J~S+4J、2St+8J~S+2J+2J-St、2St+8J~S+2J+2J-St、2St+8J~S+2J+2J-St、6St+4J~S+2J+2J-St、4St+6J~S+2J+2J-St、2St+8J~S+4J、2St+8J~S+4J和4St+6J~S+4J,与目前已报道的八倍体小偃麦均有所不同。研究结果可为这些新型八倍体小偃麦的研究和有效利用提供参考依据。     

Characterization of wheat-Thinopyrum partial amphiploids for resistance to barley yellow dwarf virus

Resistance to viruses such as wheat streak mosaic virus (WSMV) and barley yellow dwarf virus(BYDV) is lacking in the primary gene pool of wheat, and therefore resistance is being introgressed from wild relatives such as Thinopyrum species. Resistance to BYDV was found in partial amphiploids (2n = 8x = 56, consisting of 42 wheat and14 alien chromosomes) obtained in hybrids between wheat and both Th. intermedium and Th.ponticum. GISH analysis revealed that the alien genomes of all but one resistant partial amphiploid were heterogeneous consisting of different ratios of St, J^s and J genome chromosomes obtained from theThinopyrum parent. Translocated chromosomes consisting of Robertsonian, interstitial and terminal translocations between the different genomes were also detected. The tissue blot immunoassay showed that partial amphiploids having resistance could be inoculated with the virus but both virus multiplication and spread were completely blocked. This revised version was published online in August 2006 with corrections to the Cover Date.     

Fluorescent in situ hybridization as an aid to introducing alien genetic variation into wheat

Summary Fluorescent in situ hybridization (FISH) has been used to assess the occurrence and frequency of wheat-alien chromosome pairing in a wheat/Thinopyrum bessarabicum hybrid and in wheat/rye hybrids with different levels of chromosome pairing by examining pollen mother cells at metaphase I of meiosis. The use of FISH to identify the presence and size of alien chromatin in a wheat background is also demonstrated.The value of FISH as an aid to the introgression of alien genetic variation into wheat is discussed.Abbreviations FISH fluorescent in situ hybridization - GISH genomic in situ hybridization - PRINS primer-induced in situ hybridization     

The Transfer of Thinopyrum-Derived Leaf-rust Resistance from Common Wheat to Durum Wheat.

The leaf rust resistance gene on chromosome 7AL of ‘Chinese Spring’ transfer no. 12 derived from Thinopyrum ponticum, was transferred to durum wheat by standard backcrossing. In ‘Agatha’ and ‘Indis’ a leaf rust resistance gene from Thinopyrum ponticum and Thinopyrum ponticum respectively, is found on a translocated segment on chromosome arm 7DL. The use of the ‘Langdon’ disomic D-chromosome substitution lines for 7A and 7B resulted in the recovery of tetraploid leaf-rust resistant lines from the crosses with ‘Agatha’ in the B₂F₁ generation. Tetraploid lines carrying the ‘Indis’ translocation segment were recovered in the B₂F₂ generation. The F₂ segregation ratios for rust resistance after selfing or back-crossing generally fitted a 1: 1 ratio indicating non-transmission of the translocation segments in the male gametes. Homozygous resistant plants were not obtained. Meiotic instability was observed in 28 chromosome B₂ F₂ derivatives of the crosses between ‘Chinese Spring’ transfer no. 12 and durum wheat.     

Characterization of a partial amphiploid between Triticum aestivum cv. Chinese Spring and Thinopyrum intermedium ssp. trichophorum

A partial amphiploid, TE-3, between Triticum aestivum cv. Chinese Spring (CS) and Thinopyrum intermedium ssp. trichophorum was characterized by cytological observation, genomic in situ hybridization (GISH), seed storage protein electrophoresis and disease resistance screening. The TE-3 plants were deeply covered with pubescence, which is characteristic of the Th. intermedium ssp. trichophorum parent. Feulgen staining of the somatic metaphases revealed that the chromosome number varied from 52 to 56. TE-3 pollen mother cells (PMCs) regularly showed two to four univalents and 25 to 27 bivalents, indicating a degree of cytological instability. Giemsa-C banding showed that the Thinopyrum chromosomes in TE-3 produced strong heterochromatin bands. GISH analysis suggested that the alien chromosomes in TE-3 consisted of eight St chromosomes, four J^s chromosomes, and two J genome chromosomes, as well as two St-J translocation chromosomes. Seeds storage proteins separated by acid polyacrylamide gel electrophoresis (APAGE) and sodium dodecyl sulphate – polyacrylamide gel electrophoresis (SDS-PAGE) showed that TE-3 expressed some of Th. intermedium ssp. trichophorum specific gliadin and glutenin bands. When inoculated with stripe rust and powdery mildew isolates, TE-3 expressed resistance derived from its Thinopyrum parent. It appears that TE-3 can be used as a donor source in wheat breeding programs to introduce novel variation for quality and disease resistance.     

A Robertsonian translocation from Thinopyrum bessarabicum into bread wheat confers high iron and zinc contents

Development of wheat–alien translocation lines has facilitated practical utilization of alien species in wheat improvement. The production of a compensating Triticum aestivum–Thinopyrum bessarabicum whole‐arm Robertsonian translocation (RobT) involving chromosomes 6D of wheat and 6E^b of Th. bessarabicum (2n = 2x = 14, E^bE^b) through the mechanism of centric breakage–fusion is reported here. An F₂ population was derived from plants double‐monosomic for chromosome 6D and 6E^b from crosses between a DS6E^b(6D) substitution line and bread wheat cultivar ‘Roushan’ (2n = 6x = 42, AABBDD) as female parent. Eighty F₂ genotypes (L1–L80) were screened for chromosome composition. Three PCR‐based Landmark Unique Gene (PLUG) markers specific to chromosomes 6D and 6E^b were used for screening the F₂ plants. One plant with a T6E^bS.6DL centric fusion (RobT) was identified. A homozygous translocation line with full fertility was recovered among F₃ families and verified with genomic in situ hybridization (GISH). Grain micronutrient analysis showed that the DS6E^b(6D) substitution line and T6E^bS.6DL stock have higher Fe and Zn contents than the recipient wheat cultivar ‘Roushan’.     

Identification of wheat–Thinopyrum intermedium 2Ai-2 ditelosomic addition and substitution lines with resistance to barley yellow dwarf virus

Among the regenerated plants derived from immature hybrid embryos of wheat–Thinopyrum intermedium disomic addition line Z6 × common wheat variety ‘Zhong8601’, a plant with a telocentric chromosome and barley yellow dwarf virus (BYDV) resistance was obtained. The telocentric chromosome paired with an entire Thinopyrum chromosome to form a heteromorphic bivalent at meiotic metaphase I. Genomic in situ hybridization showed that the telosome originated from Th. intermedium. Two ditelosomic additions and one disomic substitution were identified among the offspring of the plant. Two random amplified polymorphic DNA molecular markers were identified among 150 random primers used to detect the different arms of the alien chromosome. These might be useful for developing translocation lines with BYDV resistance.     

Molecular cytogenetic characterization of wheat-Thinopyrum ponticum translocations bearing blue-grained gene(s) induced by r-ray

Eight γ-irradiation-induced Triticum aestivum – Thinopyrum ponticum translocations conveying the blue aleurone were characterized using molecular cytogenetic approach. The size of alien chromosome segments was estimated by genomic in situ hybridization (GISH). The wheat chromosome segments involved in these translocations were clearly identified by two-color fluorescence in situ hybridization (FISH) with the probes of pAs1 and pSc119.2 (or pHvG38). Most of the detected translocations were reciprocal translocations involving wheat chromosomes 1B, 2D, 3A, 4A, 5B, 6B, 6D and 7A. This series of blue-grained wheat translocation lines would be useful in theoretical studies and wheat chromosome engineering breeding.     

Introduction of allien variation into wheat by gene recombination. I. Crosses between mono V (5B) Triticum aestivum L. and Secale cereale L. and Aegilops columnaris zhuk.

An analysis is made of meiotic pairing of hybrids whether or not deficient in the chromosome V (5B) from crosses between Mono V (5B) T. aestivum var. Chinese Spring and Secale cereale and Aegilops columnaris. Transmission rates of deficiency are not equal for both combinations. Isochromosomes are found in the hybrid plants.5B deficiency allows the pairing and recombination between the chromosomes of wheat and those of the related species. Such deficiency may be a useful wheat breeding tool to introduce alien variation into such cultivated species.Effectiveness of 5B deficiency on meiotic pairing of the hybrids, measured as the increase of the proportion of complement paired, is 36–37% and 49–50% in the hybrids involving rye and Ae. columnaris respectively. Minimum percentages of the introduction of alien variation into wheat are established for the material investigated: 2% for rye and 28% for Ae. columnaris.Methods of utilisation of the 5B deficiency in a programme of wheat breeding are discussed and outlined.     

Molecular cytogenetic analysis of a durum wheat ×Thinopyrum distichum hybrid used as a new source of resistance to Fusarium head blight in the greenhouse

Fusarium head blight (FHB, scab), caused by Fusarium graminearum Schwabe, is a serious and damaging disease of wheat. Although some hexaploid wheat lines express a good level of resistance to FHB, the resistance available in hexaploid wheat has not yet been transferred to durum wheat. A germplasm collection of Triticum durum× alien hybrid lines was tested as a potential source of resistance to FHB under controlled conditions. Their FHB reaction was evaluated in three tests against conidial suspensions of three strains of F. graminearum at the flowering stage. Two T. durum×Thinopyrum distichum hybrid lines, ‘AFR4’ and ‘AFR5′, expressed a significantly higher level of resistance to the spread of FHB than other durum‐alien hybrid lines and a resistant common wheat line ‘Nyu‐Bay’. Genomic in situ hybridization using total genomic DNA from alien grass species demonstrated that ‘AFR5’ had 13 or 14 alien genome chromosomes plus 27 or 28 wheat chromosomes, while ‘AFR4’ had 22 alien genome and 28 wheat chromosomes. All of the alien chromosomes present in these two lines belonged to the J genome. ‘AFR5’ is likely to be more useful as a source of FHB resistance than ‘AFR4’ because of its relatively normal meiotic behaviour, high fertility and fewer number of alien chromosomes. ‘AFR5’ shows good potential as a source for transferring FHB resistance gene into wheat. The development of T. durum addition lines carrying resistance genes from ‘AFR5’ is underway.     

Inheritance of traits associated with seed size in groundnut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.)

A new wheat-Thinopyrum substitution line AS1677, developed from a cross between wheat line ML-13 and wheat-Thinopyrum intermedium ssp. trichophorum partial amphiploid TE-3, was characterized by fluorescence in situ hybridization (FISH), sequential Giemsa-C banding, genomic in situ hybridization (GISH), seed storage protein electrophoresis, molecular marker analysis and disease resistance screening. Sequential Giemsa-C banding and GISH using Pseudoroegneria spicata genomic DNA as probe indicated that a pair of St-chromosomes with strong terminal bands were introduced into AS1677. FISH using pTa71 as a probe gave strong hybridization signals at the nuclear organization region and in the distal region of the short arms of the St chromosome. Moreover, FISH using the repetitive sequence pAs1 revealed that a pair of wheat 1D chromosomes was absent in accession AS1677. Seed storage proteins separated by acid polyacrylamide gel electrophoresis (APAGE) and sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) confirmed that AS1677 lacked the gliadin and glutenin bands encoded by Gli-D1 and Glu-D1, further confirming the absence of chromosome 1D. The introduced St chromosome pair belonging to homoeologous group 1 was identified by newly produced genome specific markers. AS1677 is a new 1St (1D) substitution line. When inoculated with stripe rust and powdery mildew isolates, AS1677 expressed stripe rust resistance possibly derived from its Thinopyrum parent. AS1677 can be used as a donor source for introducing novel disease resistance genes to wheat in breeding programs aided by molecular and cytogenetic markers.     

小麦-偃麦草杂种后代及小麦种质资源对纹枯病的抗性

为鉴定小麦-偃麦草杂种后代以及我国小麦品种和育种中间品系对纹枯病的抗性,并且解析偃麦草染色体与纹枯病抗性的关系,在徐州和南京两个试点,采用田间病圃法对321份普通小麦品种或品系和56份小麦-偃麦草杂种后代材料进行了纹枯病抗性鉴定。在徐州试点没有发现高抗纹枯病的种质,但是有52份材料表现中抗反应型,包括34份普通小麦材料,其中萧农8506-1、小偃81、冀植4001、农大195、徐州8913和京东3066A-3的相对抗病指数高于0.7。在南京试点,全部普通小麦材料都不抗纹枯病,只有5份小麦-偃麦草种质表现中抗反应型。部分小麦-偃麦草种质的病情指数不但显著低于感病对照品种苏麦3号和扬麦158,而且还低于抗病对照品种安农8455和宁麦9号,如小麦-中间偃麦草4Ai#2或4Ai#2S附加系、代换系和易位系材料TA3513、TA3516、TA3517和TA3519及小麦-长穗偃麦草第4部分同源群染色体代换系SS767,说明中间偃麦草4Ai#2染色体和长穗偃麦草4J染色体可能与纹枯病病情指数降低有关。基因组原位杂交分析结果表明,4Ai#2染色体属中间偃麦草的J^s基因组,而长穗偃麦草与纹枯病抗性相关的第4部分同源群染色体属J基因组。虽然纹枯病与眼斑病的发病部位和症状非常相似,但抗眼斑病基因Pch1 (Madsen)和Pch2 (Cappelle-Desprez)对纹枯病无效。     

抗黄矮病小麦种质的分子标记

应用基因组原位杂交技术分析了抗小麦黄矮病种质的遗传组成,研究表明小麦一中间但麦草部分双二倍体无芒中4(2n=56)具有40条小麦染色体、5对中间僵麦草染色体、3对小麦/中间僵麦草易位染色体,其中1对是罗伯逊氏易位染色体。结果表明无芒中4与远中5的遗传组成有明显差异,是两种不同类型的材料。抗黄矮病小麦种质F940418, T10     

Characterization of a wheat–Thinopyron intermedium substitution line with resistance to powdery mildew

Among the progenies of a hybrid between common wheat Triticum aestivum L. cv. Yannong 15 and Thinopyron intermedium, plant E99018 was identified with the chromosome number 2n = 42 and stable agronomic traits. An analysis of the metaphase chromosome pairing indicated that it formed 21 bivalents but that 2 univalents were present in the F₁ hybrid of this plant with common wheat. Resistance verification by race 15 and with mixed races of Blumeria graminis f. sp. tritici at the seedling and adult stages showed that at both stages, the plant was immune to powdery mildew. In situ hybridization with the genomic Th. intermedium and the St genome DNAs as probes and wheat DNA as a block has shown that it contained a pair of Th. intermedium chromosomes. On the basis of the hybridization pattern of the St genome probe to the critical chromosome, a conclusion was reached that this pair of chromosomes belonged to the E genome. Therefore, plant E99018 was a spontaneously formed substitution line. An analysis by 116 SSR markers indicated that the substituted wheat chromosome was 2D and the most likely substitution in E99018 is 2E(2D).     

Identifying the alien chromosomes in wheat – Leymus multicaulis derivatives using GISH and RFLP techniques

Genomic in situ hybridization (GISH) and restriction fragment length polymorphism (RFLP) were used to identify the Leymus multicaulis (XXNN, 2n = 28) chromosomes in wheat-L. muliticaulis derivatives. Fifteen lines containing L. multicaulis alien chromosomes or chromosomal fragments were identified. All alien chromosomes or fragments in these 15 lines were from the X genome and none were from the N genome. Eleven L. multicaulis disomic addition lines and four translocation-addition lines were identified with chromosome rearrangements among homoeologous groups 2, 3, 6 and 7. Only homoeologous group 1 lacked rearrangements in addition or translocation chromosomes. The results revealed that translocation in non-homoeologous chromosomes widely exists in the Triticeae and therefore it is necessary to identify the alien chromosomes (segments) in a wheat background using these combined techniques. During the course of the work, probe PSR112, was found to detect X genome addition lines involving L. multicaulischromosomes. This may prove to be a valuable probe for the identification of alien chromosomes in a wheat background. This revised version was published online in August 2006 with corrections to the Cover Date.