人工合成甘蓝型油菜早期世代基因组变异的AFLP和MSAP标记研究

为了揭示人工甘蓝型油菜早期世代遗传和表观遗传变异规律, 以A组合(大黄油菜×中花芥蓝) S₀世代、B组合(大黄油菜×中迟芥蓝) S₀和S₁世代人工甘蓝型油菜为材料, 分别利用AFLP和MSAP技术检测基因组变化及甲基化模式变化情况。结果表明, 16对引物在A组合S₀扩增到523条带, 其中4对引物扩增出9条变异带, 包括7条亲本缺失带和2条新增带, 分别占S₀总条带的1.33%和0.38%;45对引物在B组合双亲植株扩增到1093条带, 只有1对引物检测到1条父本带型在所有S₀植株中缺失, 约占S₀总条带的0.09%;在B9子代F19-1~F19-16总共扩增得到1092条带, 变异带有10条, 占总条带的0.915%, 其中包括9条缺失带和1条新增带, 9条缺失带全部位于C基因组。MSAP检测发现, B组合S₀植株中有3个位点发生了甲基化模式的改变, 全部位于A基因组, 甲基化模式改变位点占总检测位点的1.37%。研究还发现B组合S₀世代一个植株出现可遗传的花色变异, 推测该表型变异与B组合人工甘蓝型油菜中C基因组变异有关。     

偏凸-柱穗山羊草双二倍体与普通小麦不同杂种世代的染色体及性状分离特点

为探讨偏凸山羊草－柱穗山羊草双二倍体SDAU18在小麦遗传改良中的利用价值,以SDAU18和普通小麦品种烟农15及其9个杂种世代为材料,分析不同自交和回交世代染色体和性状分离的特点。结果表明,随自交和以烟农15为轮回亲本回交世代的增加,染色体数目逐渐减少,回交比自交能使后代的染色体数目更快趋近普通小麦的42条,至F₅和BC₃F₁代,染色体数目为42的植株已分别达93.9%和92.0%。与自交世代相比,回交后代减数第一分裂中期的花粉母细胞的染色体构型较为简单,回交次数过多不利于外源染色体与普通小麦染色体发生重组,一般应以回交2~3次为宜;随自交和回交世代的增进,杂种的育性提高,至F₃和BC₂F₁代育性基本稳定。在不同杂种世代可分离出具有矮秆、大穗、大粒、对白粉病、条锈病免疫或高抗及外观品质优良的变异类型,以F₃和BC₁F₁代的变异类型最丰富。     

从携带S5n基因的水稻种质中发掘San、Sbn和Scn基因

花粉不育是籼粳杂种F₁优势利用的主要障碍之一。包括S_a、S_b和S_c等至少6个基因座位内的等位基因互作会引起花粉不育,这些座位上的中性等位基因可以克服不育性。所以,发掘和利用中性等位基因具有重要意义。本文用携带S₅ⁿ的水稻种质,分别与台中65及其携带花粉不育基因的一套近等基因系杂交,组配具有单个座位互作和多个座位同时互作的杂种F₁,首先通过观察杂种F₁的花粉育性并比较相应杂种F₁育性的差异,初步判断是否具有中性等位基因,然后,采用与S_a、S_b和S_c座位紧密连锁的分子标记对F₂植株基因型的分离进行检测,并分析其分离比例的符合度,确定存在中性等位基因的真实性。结果发现在所鉴定的6份材料中有2份(灰背子和Madhukar)同时携带S_aⁿ和S_bⁿ,3份(饭毫皮、秕五升和粤泰B)携带S_bⁿ,1份(Jackson)携带S_cⁿ。这些材料同时携带可克服杂种F₁胚囊不育和花粉不育的基因,是克服籼粳杂种F₁不育性的重要基因来源。     

OsGA20ox2不同长度RNAi片段对水稻株高等农艺性状的遗传效应

利用OsGA20ox2基因序列构建不同长度的RNAi片段并导入水稻，获得不同高度的矮化植株。将这些矮化植株与野生型植株回交获得B₁F₂群体，卡方检测表明B₁F₂群体矮秆植株数和高秆植株数符合3∶1比例，表现为矮秆显性的遗传规律。对矮化植株的F₅和B₁F₂群体株高、各茎节间长度和一些主要农艺性状方差分析显示, OsGA20ox2基因的RNAi能显著缩短株高和各节间长度(P<0.05)，RNAi干扰片段越长，使植株株高和节间长度缩短程度越大，可使株高降低24~42 cm，矮化22%~39%。在同一长度的RNAi干扰片段下，倒一节节间长度平均缩短与倒二节节间长度平均缩短非常相近，倒三节和倒四节节间长度平均缩短非常相近，总的缩短程度是倒四节>倒三节>倒二节>倒一节。这种近基部节间长度缩短幅度和比例较大的特点，利于提高水稻的抗倒伏能力，同时上部节间缩短幅度和比例较小，有效地保持合理株高，不使生物产量明显降低，有利于水稻的稳产和高产。OsGA20ox2基因的RNAi不影响如千粒重、结实率、穗长等其他主要农艺性状或影响很小。     

连续回交对消除农杆菌介导转化引起水稻体细胞变异的影响

农杆菌介导的转化引起许多体细胞变异, 影响了转基因植物的农艺性状。因此, 转基因作物的培育需要大量的T₀代再生植株。在本研究中, 我们将转基因水稻株系与原始品种连续回交, 然后评价其回交后代的表现, 消除体细胞变异, 恢复转基因亲本的农艺性状。回交的供体亲本是3个转基因水稻株系, 分别带有来自于苏云金芽胞杆菌(Bt)的抗虫基因。与原始品种连续回交至BC₃F₁代, 每代BC_nF₁单株再自交两代, 同时对各个世代进行抗虫性选择。通过发芽试验获得转基因纯合的BC_nF₃株系, 在室内抗性试验中, 所有的BC_nF₃纯合株系都能杀死100%的幼虫。在田间试验中, 这些株系的单株产量明显高于供体亲本, 大部分农艺性状与原品种没有显著的差异。这些结果说明连续回交能够在很大程度上恢复转基因水稻株系的农艺性状, 从而减少转基因育种过程中所需的工作量。     

矮秆陆地棉陆矮1号主茎顶芽几种激素含量研究

 在棉花株高建成的初蕾到盛花期,研究了陆地棉矮秆品系陆矮1号与高秆对照TM-1及其杂种F₁顶芽内源激素含量变化。结果表明：各时期陆矮1号顶芽中GA₃和IAA含量均低于TM-1和F₁,盛蕾期顶芽GA₃含量分别仅为TM-1和F₁的28.37%和31.04%,初花期IAA含量分别是TM-1和F₁的61.20%和52.76%;陆矮1号和F₁顶芽Z含量最高值出现在初蕾期,随后下降,较TM-1早15 d;陆矮1号顶芽ABA含量除初蕾期与其它品种无显著差异外,其余时期均最高,在盛蕾期、初花期和盛花期分别比TM-1高26.7%、32.4%、132.5%,比F₁高129.1%、87.5%、37.4%。     

人工合成甘蓝型油菜及其亲本的甲基化变异模式分析

甘蓝型油菜作为多倍体起源和发生的历史较短, 遗传背景较为狭窄, 人工合成甘蓝型油菜可作为植物多倍化研究的优选模型, 本文以人工合成的甘蓝型油菜为材料, 通过HPLC分析发现白菜型油菜和甘蓝的甲基化率分别为8.33%和15.88%, 2个杂种株系的全基因组甲基化水平介于双亲之间(分别为10.29%和12.83%)。MSAP分析发现杂种F₁代及其亲本的甲基化水平存在明显差异(白菜型油菜<杂种F₁<甘蓝), 杂种F1代的甲基化变异(23.71%)中来自A、C基因组的变异分别占6.60%和10.16%。MSAP差异性条带的序列分析发现多倍化过程中与甲基化变化相关的基因参与了多种生物学过程, 且差异甲基化基因在人工合成甘蓝型油菜及其亲本间的表达差异与甲基化修饰模式是一致的。本研究为了解甘蓝型油菜多倍化过程中发生的表观变异奠定了基础。     

棉属野生种克劳茨基棉第7染色体上马克隆值QTL的挖掘与定位

为了深入挖掘和利用棉属野生种克劳茨基棉(Gossypium klotzschianum)的优异等位基因，构建了一个(陆地棉泗棉2号×克劳茨基棉)×泗棉2号的BC₁F₂群体，对纤维品质性状初步定位，单标记相关分析表明位于第7染色体上的SSR标记NAU1362与马克隆值表现极显著相关。进一步选择在第7染色体上含有克劳茨基棉渐渗片段的BC₁F₂单株与轮回亲本泗棉2号回交，构建BC₂F₃和BC₂F₄分离群体，通过两年的田间重复试验验证该QTL的位置与效应。结果表明，该QTL (qFMIC-7-1)在BC₂F₃、BC₂F₄世代均被检测到，位于相同的标记区间，分别可以解释9.0%、8.8%的表型变异，增效基因来源于野生种克劳茨基棉，与BC₁F₂群体定位结果基本一致。同时在第7染色体上检测到另一马克隆值QTL (qFMIC-7-2)，同样在BC₂F₃、BC₂F₄两个世代均能够被检测到，分别可以解释3.7%、4.7%的表型变异，但增效基因均来源于泗棉2号。     

两个杂交棉F1、F2代及亲本冠层结构与物质生产特征

以杂交棉石杂2号和新陆早43的F₁、F₂代及亲本NT2、H2、4-14为试材,通过测定不同生育时期各材料叶面积指数(LAI)、叶倾角(MTA)、冠层光截获率等指标,分析了各指标变化对群体光合物质生产的影响。结果表明,2个杂交棉F₁代LAI具有超亲优势,冠层光截获率具有中亲优势; LAI和冠层光截获率具有明显的母系遗传特性,而MTA受到父本的显著影响。F₂代冠层结构主要受F₁代相关指标和衰退率的影响,LAI中亲优势减小了衰退率; 杂交棉F₁代光合物质积累主要受亲本参数和超亲优势的影响,F₂代主要受F₁代参数的影响。杂交棉光合物质积累最大增长速率和直线增长期开始时间较晚,直线增长期及活跃增长期较长,最终积累量和最大增长速率较高。杂交棉F₁代具有明显的光合生产和产量优势,F₂代具有一定的产量优势。以选择具有优化冠层结构的亲本为基础,组配具有较大MTA的父本和较大LAI的母本,有利于改善杂交棉光合性能,提高群体光能利用率,进一步挖掘产量潜力,为杂交棉高光效组合的选育及提高F₂代应用提供理论依据。     

寒地多年生小麦的选育与细胞遗传学分析

通过杂交方法获得八倍体小偃麦与中间偃麦草杂种后代，对该杂交后代进行了形态学观察和细胞遗传学分析。杂交当代结实率为10%~39%；F₁表现为两亲中间型，多年生，抗小麦多种病害，生长的第2和第3年结少量种子，结实率为2%~3%；F₂分离复杂，出现八倍体小偃麦类型和中间偃麦草类型的多年生材料；F₃和F₄代出现一些普通小麦类型的多年生小麦,表现多分蘖、多小穗、抗病、抗寒。F₁根尖减数分裂中发现49条染色体，在减数分裂中期I形成14~17个二价体和4~21个单价体；而F₂和F₃代减数分裂时形成14~21个二价体和9~17个单价体。杂种后代结实率逐代恢复。F₁植株已在田间自然条件下生长5年。从F₄代中获得了4个植株高大(140 cm)、分蘖丰富(60个以上)、小穗多(25~30个)的饲草型多年生小麦株系，它们不仅具有良好的刈割再生能力，而且兼抗多种病害，抗寒性好，草质与中间偃麦草相似。还获得了一些普通小麦类型的多年生株系，有待进一步改良。这些结果为多年生小麦的遗传研究和利用提供了信息和材料基础。     

节节麦-黑麦双二倍体的染色体C-带分析

通过对节节麦-黑麦双二倍体及其亲本节节麦、黑麦的C-分带研究,表明节节麦-黑麦双二倍体含有其双亲完整的染色体组,其C-带带型与亲本的C-带带型基本一致,从细胞学水平上证实了该种质是人工合成的一种新的异源多倍体。     

Fertility restoration of hybrids between Solanum melongena L. and S. aethiopicum L. Gilo Group by chromosome doubling and cytoplasmic effect on pollen fertility

Reproductive fertility traits were studied in the reciprocal hybrids of the eggplant(Solanum melongena L.) and S. aethiopicum L. Gilo Group, and in synthetic amphidiploids to discover whether fertility in these reciprocal hybrids was restored by chromosome doubling. Isozyme and RAPD analyses confirmed hybridity of the hybrids and amphidiploids. Analyses of chloroplast and mitochondrial DNAs confirmed that the cytoplasm of each of the hybrids and amphidiploids was from the maternal parent. Pollen sterility of S. melongena × S. aethiopicum Gilo Group [F₁ (Mel × Aet)] was restored by chromosome doubling, while the reciprocal hybrid S. aethiopicum Gilo Group ×S. melongena [F₁ (Aet × Mel)]and its amphidiploid did not produce any pollen grains; their microspores degenerated without being released from tetrads. Hence the cytoplasm of S. aethiopicum Gilo Group seems to beresponsible for their pollen-non-formation type sterility of the hybrid. Both the F₁ hybrids did not set any fruits by either selfing or backcrossing, while their amphidiploids set fruits after pollinating with pollen from the amphidiploid of F₁ (Mel × Aet). Seeds obtained from both the amphidiploids germinated normally. Chromosome doubling has been effective in restoring fertility of the hybrids. This revised version was published online in July 2006 with corrections to the Cover Date.     

Intergeneric Hybrids between Aegilops squarrosa and Secale cereale and their Meiotic Chromosome Behaviour

Three hybrid plants with different combinations of D and R genomes were produced from crosses between Aegilops squarrosa (2×: and 4×) and Secale cereale (2× and 4×) using embryo culture. Production frequencies of mature hybrids, having the genomes DR, DDR and DDRR, were 2.0%, 5.2% and 2.2 %, respectively, of florets pollinated. Amphidiploids were obtained directly from the cross between tetraploid parents. The majority of their morphological features were intermediate between those of the parents but, as with rye, the rachis was tough. Diploid and triploid hybrids were completely seed-sterile, whereas the amphidiploid had an average self-fertility of 4.5 % with a range of 0 to 45 %. The somatic chromosome numbers of diploids, triploids and amphidiploids were 2n = 14, 21 and 28, respectively. At meiotic metaphase I, the mean chromosome associations were 0.01 III + 0.26 II + 13.4 I in diploids, 6.96 II + 7.1 I in triploids, and 10.5 II + 7.9 I in amphidiploids. In diploids, pairing between D and R chromosomes was observed in the form of end-to-end types without chiasmata. Homologous pairing between D chromosomes was predominant in plants having two sets of D genomes. Amphidiploids showed a diploid-like chromosome behaviour. Homoeologous pairing was not observed in either triploids or amphidiploids.     

Morphology,cytology and forage potential of Pennisetum americanum (L.) K. Schum. x P. Purpureum Schum. amphidiploids

Summary F₁ hybrids between short-day photoperiod- sensitive maiwa bulrush millet, Pennisetum americanum (2n=14), and elephant grass, P. purpureum (2n=28), are triploid (2n=21), sterile, and can only be propagated vegetatively. Fertile amphidiploids, (2n=42, with 21 bivalents) were produced by colchicine treatment. Good seed set was obtained upon selfing the amphidiploids and the progeny exhibited variable pollen fertility and bivalent univalent and multivalent formation.Reciprocal crosses between elephant grass and the colchicine-induced amphidiploid failed to produce seeds. The maiwa millet x amphidiploid cross resulted in poor seed set and the reciprocal failed. Maiwa x amphidiploid hybrids were pollen sterile with 2n=21, rather than the expected 2n=28, indicating chromosome elimination.Morphologically, the amphidiploids and maiwa x amphidiploid hybrids closely resembled elephant grass and the F₁ hybrid. The amphidiploids showed no superiority over the F₁ hybrid and elephant grass in either forage yield or quality but one maiwa x amphidiploid genotype surpassed a promising local elephant grass selection in forage dry matter yield and flowered for a relatively short period.     

Amphidiploids between Cyclamen persicum Mill. and C. purpurascens Mill. induced by treating ovules with colchicine in vitro and sesquidiploids between the amphidiploid and the parental species induced by conventional crosses

Summary We cultured colchicine-treated hybrid ovules in vitro to produce fertile amphidiploids of C. persicum (2n=2x=48. referred to as AA) × C. purpurascens (2n=2x=34, referred to as BB). Seedlings and mature plants were obtained from the ovules without colchicine and those exposed to 50 mg/l colchicine for 5, 10 and 15 days, whereas they were not obtained from the ovules exposed to 50 mg/l colchicine for 20 days and 500 mg/l for 5, 10, 15 and 20 days. Although 8 mature hybrids derived from the ovules without colchicine produced a few fertile pollen grains, they failed to produce viable seeds by self-fertilization. The hybrids had 41 somatic chromosomes. Four and 3 mature plants were derived from ovules exposed to 50 mg/l colchicine for 10 and 15 days, respectively. One each among 4 and 3 mature plants showed a high frequency of pollen grain fertility, produced several seeds by self-fertilization, and had 82 somatic chromosomes which is twice the number of hybrid chromosomes (2n=41, AB). These findings indicated that these plants are amphidiploids (2n=82, AABB) between C. persicum and C. purpurascens. Three and 2 viable seeds were derived by the conventional crosses of diploid C. persicum × the amphidiploid and the amphidiploid × C. purpurascens, respectively. Flowering plants that developed from the seeds of diploid C. persicum × the amphidiploid were barely fertile and had 65 somatic chromosomes (2n=65, AAB), whereas those that developed from the seeds of the amphidiploid × C. purpurascens were barely fertile and had 58 somatic chromosomes (2n=58, ABB). The somatic chromosomes indicated that these plants are probably sesquidiploids between the amphidiploid and either C. persicum or C. purpurascens. The interspecific cross-breeding of cyclamen using the amphidiploids and the sesquidiploids is discussed.     

Fertility and segregation of the amphidiploid Phaseolus vulgaris L. x P. coccineus L. and its behaviour in backcrosses

Summary Amphidiploidy was induced by colchicine treatment of cuttings from the F₁ interspecific hybrid Phaseolus vulgaris L. x Ph. coccineus L.. Pollen stainability rose from about 50% in the raw amphidiploid to 76% in some C₅ individuals; similar improvement in seed fertility was also observed. It is suggested that both genic and chromosomal factors are implicated in the control of fertility in the amphidiploid.The amphidiploid is apparently unable to cross successfully with either parental species; difference in ploidy level is apparently an effective isolating mechanism between the amphidiploid and its parents.     

小麦D基因组产量性状QTL定位

粗山羊草是普通小麦的D染色体组供体,为了寻找粗山羊草中对小麦产量性状遗传改良有益的基因,通过对四倍体硬粒小麦与粗山羊草杂交合成的双二倍体Am6-1和普通小麦品种Ph85-16的回交一代进行产量性状变异特点分析,发现粗山羊草的D组染色体对小麦的产量性状具有显著影响,千粒重、穗长、穗粒数和每穗小穗数明显高于Ph85-16;同时利用130对SSR引物对几个与产量性状相关的QTL位点进行了定位,初步寻找到4个主效QTL,它们分别为与穗长相关的QSl．sdau-5D,其贡献率为31．58％,与株高相关的QPh．sdau-1D,其贡献率为25．38％,与穗粒数相关的QGs．sdau-5D,其贡献率为44.65％,与千粒重相关的QTgw．sdau-3D,其贡献率为61．62％。     

萝卜-芥蓝异源四倍体的合成及GISH分析

通过萝卜(Raphanus sativus L.,2n=18,RR)与白花芥蓝(Brassica alboglabra Bailey,2n=18, CC)杂交,F1经秋水仙碱加倍合成萝卜-芥蓝异源四倍体（Raphanobrassica, 2n=36, RRCC）。经F₄~F₁₀代连续育性选择,F₁₀单株种子产量达32.3 g,每角粒数达14.9。基因组原位杂交显示F₁₀减数分裂行为类似于二倍体物种,表明该异源四倍体的细胞学行为已经稳定。育性观察表明,可育花粉足够各代生产种子,但低世代杂种出现高频瘪粒种子,胚珠败孕可能是其主要原因。该萝卜-芥蓝异源四倍体可以用作向油菜（B. napus L.,2n=38,AACC）转移萝卜基因的遗传桥梁。     

The characterization of autoallotetraploid hybrids between Iris ensata Thunb. and I. laevigata Fisch.

The characteristics of autoallotetraploid hybrids obtained from the cross between Iris ensata cv. Raspberry Rimmed (4X) and amphidiploids of I. laevigata × I. ensata were examined and compared with those of their parents. The color of inner and outer perianths in the autoallotetraploids were bluish purple and similar to those of the amphidiploid parent. However, the autoallotetraploids exhibited low pollen fertility. In addition, the autoallotetraploids were characterized by 17 or 19 anthocyanins and had high resemblance to their parents in the anthocyanin expression. Among these anthocyanins, malvidin 3RGac5G and petunidin 3RGac5G were regarded as major anthocyanins in the autoallotetraploids and their parents, but the differences in the ratios of malvidin 3RGac5G:petunidin 3RGac5G between the autoallotetraploids and their parents were ca. 2:1 for the former and ca. 1:1 for the latter. No viable hybrid seeds were obtained from the reciprocal crosses between I. ensata (2X and 4X) and the autoallotetraploids. Finally, the interspecific cross-breeding of I. ensata using the autoallotetraploids is discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Chromosome associations and crossability with Iris ensata Thunb. in induced amphidiploids of I. laevigata Fisch. × I. ensata

Summary The chromosome associations of amphidiploids of I. laevigata × I. ensata were analysed and compared with those of the parental species and F₁ hybrids of I. laevigata × I. ensata. The F₁ hybrids showed partial chromosome associations. Their mean chromosome association per cell was 20.73I+3.63II, although the mean chromosome association per cell in the parental species was 0.09I+15.96II for I. laevigata and 0.03I+11.98II for I. ensata, respecively. In contrast, the normal association (28II) was partially restored in the amphidiploids. Their mean chromosome association per cell was 1.93I+26.48II+0.28III+0.03IV+0.03V. In this study, moreover, the crossability between I. ensata (2X and 4X) and the amphidiploids and between I. laevigata and the amphiliploids was examined. No hybrid plants were obtained from both reciprocal crosses between I. ensata (2X) and the amphidiploids and between I. laevigata and the amphidiploids. Only the cross of I. ensata (4X) × the amphidiploids in the reciprocal crosses produced hybrid plants. The observation of their somatic chromosome numbers indicates that these are true hybrid plants between autotetraploid I. ensata and the amphidiploids, and such plants can be called autoallotetraploids between I. ensata and I. laevigata. The interspecific cross-breeding of I. ensata using the autoallotetraploids is discussed.