黄瓜属不同倍性异源多倍体的形态及生理特性分析

以黄瓜属3种不同倍性异源多倍体为试验材料,比较分析它们的形态和生理特性与基因组剂量的关系,为进一步研究黄瓜属基因组剂量效应、探讨植物多倍体进化机理奠定基础。结果表明:(1)黄瓜属异源四倍体与种间杂种F1相比,其叶片厚度、主蔓直径等性状随基因组剂量的增加而增大,而果实大小、主蔓节间长以及果瘤果刺的大小随基因组剂量的增加而减小。(2)在异源三倍体中,叶片厚度和主蔓直径等表型性状也表现出一定的基因组剂量效应。(3)基因组剂量的变化会引起黄瓜属异源多倍体中叶绿素含量、POD活性以及IAAi、PA和ZR等内源激素的变化。     

栽培黄瓜与野黄瓜正反杂交的几种同工酶分析

运用天冬氨酸转氨酶(AAT)、苹果酸脱氢酶(MDH)以及酯酶(EST)3种同工酶对栽培黄瓜"长春密刺"Cucumis sativus cv. Changchunmici (2n=14)与野黄瓜C. hystrix (2n=24)的正反交种间杂种F1 (正交: 野黄瓜×栽培黄瓜"长春密刺", 反交: 栽培黄瓜"长春密刺"×野黄瓜)及其双亲进行鉴定和比较研究。结果表明: 正反交种间杂种F1主要表现为双亲酶带的互补, 同时还形成4个杂合带(Aat-1-94、Aat-2-104、Mdh-3-102和Est-5-102)。上述3种酶均能准确地鉴定种间杂种的真实性。研究还发现正反交杂种F1的AAT和MDH的酶谱分别在酶带数目和强弱上表现出一定的差异, 进一步证实了野黄瓜与栽培黄瓜杂交存在正反交差异。     

Cucumis属3种不同倍性种间杂交后代的同工酶分析

以3种不同倍性的Cucum is属种间杂交后代[异源四倍体C.hy tivus(2n=4x=38,HHCC)、异源三倍体(2n=3x=26,HCC)和正反交种间杂种F1(2n=2x=19,HC/CH)]及其双亲为试材,比较研究了过氧化物酶(POD)和酯酶(EST)同工酶在不同器官中的酶谱表达特性.结果表明,花蕾中的POD和EST同工酶谱带都比叶片中的丰富,表现出明显的组织特异性.在相同器官的同一酶系统中,3种不同倍性种间杂交后代的酶谱基本一致,主要表现为互补双亲的酶带,同时出现了双亲所没有的酶带(POD4c和EST3b等),表明远缘杂交扩大了黄瓜的遗传基础.此外,在幼叶和花蕾的POD同工酶中,大部分酶带活性随染色体倍性增加而减弱,表明基因剂量与POD同工酶酶谱的表达呈负相关.     

[AAGG]异源四倍体新棉种的核型及同功酶分析

本文对山西农业大学棉花育种组创育的[AAGG]异源四倍体新棉种及其二倍体亲本亚洲棉(G．arboreum)、比克氏棉(G．bickii)进行了核型分析,并对[AG]异源二倍体和[AAGG]异源四倍体的过氧化物同功酶进行了分析。研究结果：异源四倍体[AAGG]的体细胞染色体数目为2n=4x=52,表现为双亲染色体数目之和,其核型公式为2n=4x=52=50m(6SAT) 2sm,按stebbins的核型分析原则,异源四倍体[AAGG]属1A型。过氧化物酶同功酶分析表明：[AG]棉种酶谱表现为双亲不完全互补型。并具有新式酶带。因此过氧化物酶同功酶可以作为鉴别远缘杂种的生物技术之一。     

黄瓜同源三倍体创制及减数分裂行为观察

采用常规杂交法研究黄瓜二、四倍体杂交过程中亲本育性、授粉组合及亲本基因型对杂交结实率的影响,并利用减数分裂制片法对获得的黄瓜同源三倍体进行了花粉母细胞减数分裂行为的观察.结果显示:(1)同源四倍体白交结实率比较低(13.0%～14.5%),可能与其花药内所包含的正常花粉粒比例小及花粉管萌发长度较短有关.(2)二、四倍体杂交组合的结实率很低(0.26%～0.02%),但在两种配组方式之间存在着明显差异,即以同源四倍体黄瓜为父本和二倍体黄瓜为母本的杂交结实率比较高,反之则杂交结实率比较低.(3)在二、四倍体杂交过程中,二、四倍体的基因型对杂交结实率的影响较大,以杂交双亲同属一个基因型的杂交效果较好.(4)同源三倍体花粉母细胞减数分裂过程与二倍体基本相同,但存在较高频率的染色体异常行为:中期I染色体构型复杂,在大多数花粉母细胞中可观察到单价体、二价体、三价体的存在;中期Ⅰ和Ⅱ有少数染色体游离于赤道板外;后期I和后期Ⅱ常出现落后染色体、染色体桥及细胞分裂不同步现象,其最终结果导致了不正常四分体和不可育配子的形成.(5)同源三倍体花粉粒的平均可染率和萌发率分别为18.8%和11.3%.研究结果表明,黄瓜二、四倍体正反交能直接获得同源三倍体材料;同源三倍体花粉母细胞减数分裂异常导致不育配子的形成是引起其育性低的细胞学原因;同源三倍体的部分育性为通过同源三倍体的回交来选育初级三体系奠定了基础.     

野生南荻与芒杂种多倍体诱导研究

用不同浓度秋水仙素处理野生南荻×芒(Miscanthus lutarioriparia×Miscanthus sinensis)远缘杂交后代以诱导产生多倍体,并对变异株进行形态学和细胞学鉴定,以期获得稳定的四倍体植株并分析其生理特性。结果表明:(1)采用秋水仙素加入培养基处理法和秋水仙素溶液浸泡处理法都可获得一定频率的多倍体植株;胚性愈伤组织以0.2%秋水仙素浸泡处理48h的诱变效果较好,四倍体诱导率达8.7%;芽在0.05%秋水仙素培养基中处理15d较好,四倍体诱导率达10.6%;生根苗在0.1%秋水仙素培养基中处理10d较好,四倍体诱导率达11.1%。(2)经体细胞染色体计数,加倍植株染色体数为2n=4x=76,对照植株的染色体数目为2n=2x=38。(3)生长2年的多倍体植株形态、叶片大小、茎粗、茎壁厚、节间等性状表现出巨大性和超亲优势。     

异源四倍体鲫鲤及其原始父母本细胞周期蛋白B基因克隆与分子遗传分析

细胞周期蛋白(cyclin)B是真核细胞周期运转中调控G2期至M期转化的关键因子.本实验根据斑马鱼细胞周期蛋白 B1基因的剪切方式,设计3对特异于金鱼和银鲫细胞周期蛋白B基因外显子区兼并引物,首次扩增出异源四倍体鲫鲤及其原始亲本红鲫和鲤鱼细胞周期蛋白B基因2条大小分别约为2.4 kb和2.1 kb的片段.测序及比对分析表明：这3种鱼的细胞周期蛋白B基因2个片段均包含8个外显子和7个内含子.内含子剪切位点符合GT/AG规则,推测细胞周期蛋白B基因2个片段可能是细胞周期蛋白B基因的2种存在形式.异源四倍体鲫鲤与其原始父母本细胞周期蛋白B基因片段序列的比较结果表明：无论是外显子区还是内含子区,异源四倍体鲫鲤与其原始亲本都具有较高的遗传相似性,在1 025 bp的外显子序列中相同的碱基位点数达963个,为异源四倍体鲫鲤来源于红鲫和鲤鱼提供了分子证据;同时,异源四倍体鲫鲤与其原始亲本差异碱基位点的存在又表明这一独特的多倍体物种与其原始亲本存在着进化上的变异.此外,还分别以细胞周期蛋白B基因外显子和内含子序列构建了包括异源四倍体鲫鲤及其原始父母本在内的系统进化树.结果初步表明：对于亲缘关系较近的物种,用外显子和内含子序列构建的系统进化树与传统的物种进化树一致;而对于亲缘关系较远的物种,用内含子序列构建的进化树与传统的物种进化顺序存在较大差异.     

在六倍体小麦的异源多倍化早期微卫星侧翼序列迅速发生了改变

在异源多倍体形成的早期, DNA序列和基因的表达迅速发生了改变. 以异源六倍体小麦为例, 比较了四倍体小麦与节节麦合成六倍体小麦前后, 位于普通小麦D染色体组不同染色体臂上的特异性引物揭示的微卫星位点变化特点. 结果表明, 从四倍体小麦与节节麦杂交, 将A, B与D染色体组结合在一起并加倍得到AABBDD的六倍体小麦这一“剧烈事件”中: (ⅰ) 微卫星的侧翼序列发生了变化, 导致出现了供体物种没有的新带纹或供体物种的带纹消失, 其中, 供体物种的带纹消失是主要的. (ⅱ) 供体物种的带纹消失不是随机的, 而是四倍体小麦消失频率远高于节节麦的频率, 即发生在A, B染色体组的消失频率比发生在D染色体组的频率高得多. (ⅲ) 微卫星侧翼序列的变化在多倍化的早期(F₁代或S₁代)就开始发生. 由此看来, 微卫星两边的侧翼区域在多倍化过程中很活跃, 是容易发生变化的区域. 微卫星的生物学功能可能与多倍体进化过程有关, 微卫星两边的侧翼区域在多倍化过程的早期迅速发生有方向性的改变可能有利于新形成异源多倍体的迅速进化, 从而使不同染色体组在遗传上迅速达到协调.     

远缘杂交形成的二倍体鱼和多倍体鱼生殖细胞染色体研究

本文采用性腺染色体制片及组织学切片方法,系统地研究了不同发育时期的鲫鲤杂交第二代(F2) (2n=100)、异源四倍体鲫鲤(4n=200)、三倍体鲫鱼(3n=150))、雌核发育二倍体鲫鲤第二代(G2)(2n=100)及鲤鱼(Cypninus carpio L)(2n=100)(对照组)生殖细胞的染色体特征.研究结果表明,对照组中鲤鱼精原细胞染色体数与体细胞染色体数一致,为二倍体精原细胞(2n=100),而远缘杂交形成的二倍体鱼和多倍体鱼的生殖细胞中则观察到明显的染色体数加倍现象,其中,鲫鲤杂交第二代(F2)精巢生殖细胞染色体数加倍现象特别丰富,占检测的染色体分裂相的21.6%,为其产生不减半的二倍体配子提供了直接的细胞学证据,同时也说明远缘杂交是导致生殖细胞染色体数加倍的一个重要因素.该研究在探讨多倍体鱼的发生及鱼类遗传育种方面具有重要意义.     

不同倍性黄瓜遗传差异的AFLP分析

采用AFLP(Amplified fragment length polymorphism)技术对黄瓜(Cucumis sativusL.)品种‘津绿4号’的单倍体、二倍体和四倍体进行基因组DNA多态性比较。结果表明:(1)从35对引物扩增获得2 188条60~500 bp的条带,多态性位点仅有20个,占0.92%,其中22对引物组合扩增的不同倍性材料的AFLP指纹没有明显差异;(2)在多态性位点表现中,以二倍体的条带存在,单倍体和(或)四倍体的条带丢失为主,在四倍体中扩增出1条特异带;(3)与相应的二倍体相比,单倍体和四倍体有特异片段的消失和增加。     

Allopolyploidy-Induced Rapid Genome Evolution in the Wheat (Aegilops–Triticum) Group

To better understand genetic events that accompany allopolyploid formation, we studied the rate and time of elimination of eight DNA sequences in F1 hybrids and newly formed allopolyploids of Aegilops and Triticum. In total, 35 interspecific and intergeneric F1 hybrids and 22 derived allopolyploids were analyzed and compared with their direct parental plants. The studied sequences exist in all the diploid species of the Triticeae but occur in only one genome, either in one homologous pair (chromosome-specific sequences [CSSs]) or in several pairs of the same genome (genome-specific sequences [GSSs]), in the polyploid wheats. It was found that rapid elimination of CSSs and GSSs is a general phenomenon in newly synthesized allopolyploids. Elimination of GSSs was already initiated in F1 plants and was completed in the second or third allopolyploid generation, whereas elimination of CSSs started in the first allopolyploid generation and was completed in the second or third generation. Sequence elimination started earlier in allopolyploids whose genome constitution was analogous to natural polyploids compared with allopolyploids that do not occur in nature. Elimination is a nonrandom and reproducible event whose direction was determined by the genomic combination of the hybrid or the allopolyploid. It was not affected by the genotype of the parental plants, by their cytoplasm, or by the ploidy level, and it did not result from intergenomic recombination. Allopolyploidy-induced sequence elimination occurred in a sizable fraction of the genome and in sequences that were apparently noncoding. This finding suggests a role in augmenting the differentiation of homoeologous chromosomes at the polyploid level, thereby providing the physical basis for the diploid-like meiotic behavior of newly formed allopolyploids. In our view, this rapid genome adjustment may have contributed to the successful establishment of newly formed allopolyploids as new species.     

Allopolyploidy-induced rapid genome evolution in the wheat (Aegilops-Triticum) group

To better understand genetic events that accompany allopolyploid formation, we studied the rate and time of elimination of eight DNA sequences in F1 hybrids and newly formed allopolyploids of Aegilops and TRITICUM: In total, 35 interspecific and intergeneric F1 hybrids and 22 derived allopolyploids were analyzed and compared with their direct parental plants. The studied sequences exist in all the diploid species of the Triticeae but occur in only one genome, either in one homologous pair (chromosome-specific sequences [CSSs]) or in several pairs of the same genome (genome-specific sequences [GSSs]), in the polyploid wheats. It was found that rapid elimination of CSSs and GSSs is a general phenomenon in newly synthesized allopolyploids. Elimination of GSSs was already initiated in F1 plants and was completed in the second or third allopolyploid generation, whereas elimination of CSSs started in the first allopolyploid generation and was completed in the second or third generation. Sequence elimination started earlier in allopolyploids whose genome constitution was analogous to natural polyploids compared with allopolyploids that do not occur in nature. Elimination is a nonrandom and reproducible event whose direction was determined by the genomic combination of the hybrid or the allopolyploid. It was not affected by the genotype of the parental plants, by their cytoplasm, or by the ploidy level, and it did not result from intergenomic recombination. Allopolyploidy-induced sequence elimination occurred in a sizable fraction of the genome and in sequences that were apparently noncoding. This finding suggests a role in augmenting the differentiation of homoeologous chromosomes at the polyploid level, thereby providing the physical basis for the diploid-like meiotic behavior of newly formed allopolyploids. In our view, this rapid genome adjustment may have contributed to the successful establishment of newly formed allopolyploids as new species.     

序列消除与异源多倍体植物基因组的进化

经杂交后多倍化形成的异源多倍体植物,被认为在其形成的早期阶段经历了DNA序列消除过程。发生消除的序列既涉及到高拷贝的序列也有低拷贝的序列,而且大多数情况下倾向于消除来自其中一个亲本的序列。序列消除的模式因基因组组成和物种的不同而有差异,并且可能受到细胞质的影响。尽管序列消除的分子机制还不是很清楚,但很多证据已表明非同源染色体之间的互作不是主要的原因。目前认为,序列消除增加了非同源染色体之间的差异,为多倍化后在减数分裂过程中快速恢复二倍化的染色体配对模式提供了物质基础,这样更有利于多倍体在自然界快速稳定。     

Paternal inheritance of egg traits in mice: a case of genomic imprinting

Eggs from reciprocal hybrids between the C57BL/6By and BALB/cBy strains were tested for their susceptibility to attack by hyaluronidase and pronase. There were significant reciprocal differences between the F1 females in the responses of their unfertilized eggs to both enzymes. The F1 hybrids from BALB mothers showed the increased susceptibility characteristic of C57BL whilst the F1 hybrids with C57BL mothers were more resistant to both enzymes, like BALB mice. Eggs from the four kinds of reciprocal F2 hybrid females also showed patroclinous patterns of susceptibility. A patroclinous difference was found between reciprocal crosses of the CXBD and CXBE recombinant inbred strains but not in crosses between recombinant inbred strains with similar phenotypes. Cross fostering did not alter the phenotypes of the C57BL and BALB females or those of their reciprocal F1 hybrids. The findings are interpreted in terms of differential genomic imprinting of paternally inherited information. The possible general usefulness of patroclinous differences between reciprocal F1 females in revealing differences in imprinting is noted.     

Desiccation resistance in interspecific Drosophila crosses. Genetic interactions and trait correlations

We used crosses between two closely related Drosophila species, Drosophila serrata and D. birchii, to examine the genetic basis of desiccation resistance and correlations between resistance, physiological traits, and life-history traits. D. serrata is more resistant to desiccation than D. birchii, and this may help to explain the broader geographical range of the former species. A comparison of F2's from reciprocal crosses indicated higher resistance levels when F2's originated from D. birchii mothers compared to D. serrata mothers. However, backcrosses had a resistance level similar to that of the parental species, suggesting an interaction between X-linked effects in D. serrata that reduce resistance and autosomal effects that increase resistance. Reciprocal differences persisted in hybrid lines set up from the different reciprocal crosses and tested at later generations. Increased desiccation resistance was associated with an increased body size in two sets of hybrid lines and in half-sib groups set up from the F4's after crossing the two species, but size associations were inconsistent in the F2's. None of the crosses provided evidence for a positive association between desiccation resistance and glycogen levels, or evidence for a tradeoff between desiccation resistance and early fecundity. However, fecundity was positively correlated with body size at both the genetic and phenotypic levels. This study illustrates how interspecific crosses may provide information on genetic interactions between traits following adaptive divergence, as well as on the genetic basis of the traits.     

Asymmetric postmating isolation: Darwin's corollary to Haldane's rule

Asymmetric postmating isolation, where reciprocal interspecific crosses produce different levels of fertilization success or hybrid sterility/inviability, is very common. Darwin emphasized its pervasiveness in plants, but it occurs in all taxa assayed. This asymmetry often results from Dobzhansky-Muller incompatibilities (DMIs) involving uniparentally inherited genetic factors (e.g., gametophyte-sporophyte interactions in plants or cytoplasmic-nuclear interactions). Typically, unidirectional (U) DMIs act simultaneously with bidirectional (B) DMIs between autosomal loci that affect reciprocal crosses equally. We model both classes of two-locus DMIs to make quantitative and qualitative predictions concerning patterns of isolation asymmetry in parental species crosses and in the hybrid F(1) generation. First, we find conditions that produce expected differences. Second, we present a stochastic analysis of DMI accumulation to predict probable levels of asymmetry as divergence time increases. We find that systematic interspecific differences in relative rates of evolution for autosomal vs. nonautosomal loci can lead to different expected F(1) fitnesses from reciprocal crosses, but asymmetries are more simply explained by stochastic differences in the accumulation of U DMIs. The magnitude of asymmetry depends primarily on the cumulative effects of U vs. B DMIs (which depend on heterozygous effects of DMIs), the average number of DMIs required to produce complete reproductive isolation (more asymmetry occurs when fewer DMIs are required), and the shape of the function describing how fitness declines as DMIs accumulate. Comparing our predictions to data from diverse taxa indicates that unidirectional DMIs, specifically involving sex chromosomes, cytoplasmic elements, and maternal effects, are likely to play an important role in postmating isolation.     

Genetically determined transient edema found in the WB/ReJ mouse strain in a teratogenic survey with acetazolamide

A continuing survey of the genetic variability of different mouse strains to acetazolamide teratogenesis demonstrated the WB/ReJ strain expresses a high frequency of induced subcutaneous edema in day 15 fetuses. In treated WB/ReJ fetuses, the probability of expression of edema is independent of the expression of forelimb ectrodactyly and, with the dosage regime, there is no significant increase in acetazolamide-induced resorption. It was surprising to find a high frequency of spontaneous edema on day 15 in untreated WB/ReJ fetuses. The spontaneous edema is a transient trait with maximum expression (56%) on day 14, and it is resolved by day 17 without apparent consequence to the survival of previously affected fetuses. There is no sex dimorphism in the liability to express the transient edema. Preliminary genetic crosses to investigate the spontaneous edema were made between WB/ReJ and the C57BL/6J strain, which historically had not be observed to express spontaneous edema. A low frequency of spontaneous edema was observed on day 14 in both C57BL/6J and the reciprocal F1 fetuses. The trait is not additive because there is dominance deviation of the BC1 fetuses in the direction of the F1 fetuses. The data were fitted to a threshold model suggesting that the developmental liability to express the difference in transient edema is determined by more than one gene, but the data can be interpreted by a minimum of two loci with duplicate epistasis. The observed differences in frequencies of edema suggest the genetic model can be tested with relatively few test crosses.     

Genetics of Glucocorticoid Receptor Levels in Recombinant Inbred Lines of Mice

Hepatic glucocorticoid receptor binding activity was measured in A/J, C57BL/6J, their F1 reciprocal crosses and their F1 recombinant inbred (RI) lines. The glucocorticoid binding capacity was measured in Hepes buffer and Hepes buffer plus dithiothreitol (DTT). The A/J parental strain showed higher levels, and a greater increase of glucocorticoid binding in the presence of DTT, than did the C57BL/6J strain. The response of binding in the presence of DTT to that without DTT was expressed as a ratio. The levels and distribution of these measurements among the RI lines and F1 reciprocal crosses suggested that there was a maternal effect on glucocorticoid receptor binding capacity. The data on RI lines suggested epistatic interactions, but could fit a two-gene model. beta 2-Microglobulin, beta-glucuronidase and H-2 (located on chromosomes 2, 5, and 17, respectively) were chosen to analyze any association to glucocorticoid receptor binding, because they have been considered to be related to glucocorticoid-induced cleft palate or glucocorticoid receptors. No significant associations were found.     

Genome evolution of allopolyploids: a process of cytological and genetic diploidization

Allopolyploidy is a prominent mode of speciation in higher plants. Due to the coexistence of closely related genomes, a successful allopolyploid must have the ability to invoke and maintain diploid-like behavior, both cytologically and genetically. Recent studies on natural and synthetic allopolyploids have raised many discrepancies. Most species have displayed non-Mendelian behavior in the allopolyploids, but others have not. Some species have demonstrated rapid genome changes following allopolyploid formation, while others have conserved progenitor genomes. Some have displayed directed, non-random genome changes, whereas others have shown random changes. Some of the genomic changes have appeared in the F1 hybrids, which have been attributed to the union of gametes from different progenitors, while other changes have occurred during or after genome doubling. Although these observations provide significant novel insights into the evolution of allopolyploids, the overall mechanisms of the event are still elusive. It appears that both genetic and epigenetic operations are involved in the diploidization process of allopolyploids. Overall, genetic and epigenetic variations are often associated with the activities of repetitive sequences and transposon elements. Specifically, genomic sequence elimination and chromosome rearrangement are probably the major forces guiding cytological diploidization. Gene non-functionalization, sub-functionalization, neo-functionalization, as well as other kinds of epigenetic modifications, are likely the leading factors promoting genetic diploidization.