植物多倍体研究的回顾与展望

多倍化是促进植物进化的重要力量。多倍体主要是通过未减数配子融合,体细胞染色体加倍以及多精受精三种方式起源的。其中,不减数配子是多倍体形成的主要机制。三倍体可能在四倍体的进化中起了重要作用。过去认为多倍体只能是进化的死胡同,现在发现很多多倍体类群都是多元起源的而不是单元起源的。当多倍体形成后,基因组中的重复基因大部分保持原有的功能,也有相当比例的基因发生基因沉默。多倍体通常表现出不存在于二倍体祖先的表型,并且超出了其祖先的分布范围,因为在多倍体中发生了很多基因表达的变化。主要从多倍体的起源、影响多倍体发生的因素及多倍体基因组的进化等方面回顾并展望多倍体的研究。     

由鲤(Cyprinus carpio)和鲫(Carassius auratus)染色体组叠加构建的 两个单性人工多倍体克隆

具有天然雌核发育的多倍体杂种鱼可防止杂种优势的分离并保持其后代的杂种优势. 由于假设诱发的多倍体鱼类的生殖模式是天然雌核发育的, 我们进行了鲤鲫杂种的多倍体诱发, 目的是描述经染色体组叠加由有性鲤鲫二倍体转化为异源三倍体及异源四倍体克隆谱系. 鲤鲫杂种产生未减数而具有两亲本染色体组杂种卵子, 未减数的雌核可与入卵的雄核融合叠加形成三倍体合子. 鲤鲫异源三倍体胚胎发育正常, 部分异源三倍体雌性个体可产生未减数的、仍保留母本的三套染色体的成熟卵子. 绝大部分鲤鲫异源人工三倍体个体的成熟卵子的雌核不与入卵的雄核融合, 具有天然雌核发育特性. 异源三倍体卵子在入卵精子的激动下由雌核发育产生全雌后代, 并形成一个单性克隆系, 后代保留异源三倍体母本的形态特征, 并靠雌核发育的生殖方式形成异源三倍体克隆系. 极少数异源三倍体个体的成熟卵子的雌核可与入卵的雄核融合, 再通过染色体组叠加形成鲤鲫异源四倍体. 所有异源四倍体的雌性产生未减数的、含有4个染色体组的成熟卵子. 异源四倍体的成熟卵子保持雌核发育特性, 在近类的精子诱发下产生单性后代, 形成一个异源四倍体单性克隆.     

人工诱导异源四倍体和倍间三倍体白鲫的红细胞观察及其相对DNA含量测定

染色体核型分析表明,异源四倍体白鲫含有两套白鲫染色体和两套红鲫染色体,新四倍体白鲫含有三套白鲫染色体和一套红鲫染色体,倍间三倍体白鲫含有二套白鲫染色体和一套红鲫染色体。为进一步阐明人工诱导多倍体的染色体倍性与细胞和细胞核及其相对DNA含量之间的相关关系,作者测量了异源四倍体、新四倍体和借间三倍体鱼红细胞及其核的大小,同时测定了它们的红细胞相对DNA含量,并与它们亲本红细胞及其精子细胞相对DNA含量进行了比较。       

植物同源多倍体育种的几个问题

四倍体变异既来自染色体数量、结构的变异以及基因突变、重组,同时也与核一质之间、合子胚与胚乳之间协调关系的变化有关。其变异率可以预料高于二倍体。纯合的四倍体是一个相对平衡的系统,继续选择是无效的。不同物种四倍体的表型变异有一定差别,经常是既有有利变异,也有不利变异。四倍体育种应取利避害,针对具体情况采用适宜的选择方法。减数分裂四倍体由2x的2n配子和经选择的4x的正常配子融合产生,避免了化学诱变的不良影响,能够较好的实现双亲优良性状的互补,也比较容易稳定。四倍体可以直接育成稳定品种,也可以进行杂种优势的利用。利用多倍体作桥梁种有利于促进种或种群间的基因交流。三倍体可能表现出强的杂种优势,也可能产生无籽果实,但并不是所有物种都能直接利用三倍体品种进行繁育。对四倍体种或二倍体种或人工四倍体种的改良选择,利用双单倍体都具重要意义。利用多倍体选育二倍体品种,进行非倍性诱变育种是多倍体育种的重要组成部分。多倍体和无融合生殖结合利用,利用多倍体创造新种质,进行遗传研究也有重要价值。     

遗传学专业术语不规范化的现象探研—I.同名异义现象

对现行的遗传学教材中存在的专业术语同名异义现象进行探研,针对Y连锁遗传、限性遗传、从性遗传、单倍体、多倍性和累加作用等专业术语存在的表达不清,相互矛盾,同名异义等问题,进行分析探讨,并提出Y连锁遗传不能与限性遗传相混淆,限性遗传和从性遗传的基因是位于常染色体上,但二者存在区别。单倍体是指具有配子染色体数目的个体,但并不等同于一倍体。建议多倍体用染色体基数来表达,如三倍体用3X表示。 累加作用是指数量性状遗传基因间的相互累加作用。     

芸薹属作物的渗入杂交与种质资源创新

通过人工合成的种间杂种或异源多倍体与栽培种连续回交与选择,可使近缘种或野生种的目标性状与基因渗入作物。本文总结了本课题组30年间所开展的芸薹属栽培种与几个亲缘关系较远的物种间的族间杂交、特异的细胞遗传学行为及种质资源创建。由于这些族间杂交中发生父本染色体的部分消除,产生的杂种具有母本栽培种的全部或大部分染色体、附加少数的外源染色体或染色体片段,经回交与自交后获得的渗入系表现父本的一些特征、遗传与表观遗传发生变化、但恢复母本的染色体数。讨论了此类亲缘关系较远的杂交在创建新种质资源方面的应用前景。     

林木三倍体育种研究进展及展望

植物三倍体具有生长迅速、叶片或花、果实硕大、新陈代谢旺盛、适应性强以及不孕等特点,尤其适用于无性繁殖林木的遗传改良.利用配子染色体加倍或四倍体与二倍体杂交等有性多倍化技术手段,增加目标树种一套基因组的全部同源基因,可以仅通过一轮次的育种过程实现多目标性状综合改良,培育出生长快、纤维长、木素低、纤维含量高以及抗逆性增强的用材林新品种,或者是生长迅速、叶片和果实等器官巨大、含胶量及药物成分含量显著提高的胶用、漆用、药用、叶用、果用新品种.在人工诱导配子染色体加倍选育三倍体时,实现有效处理时期的即时判别是提高三倍体育种效率的关键,而选配高配合力亲本则是提高三倍体育种效果的保障.随着林木三倍体育种理论和技术方法的突破及其不断向其他树种拓展,三倍体育种必将在提高林产品产量、改善品质以及增强抗逆性等方面发挥更大的作用.     

使用染色体数目符号的建议

我们感到目前使用染色体数目的符号不够准确,常常看到一些文章、小册子是这样叙述染色体数目的。 “以”表示基本染色体组的染色体数,称为单倍体,则含有二个染色体组的细胞或个体称为二倍体,用2n表示;含有三个染色体组的细胞或个体称为三倍体,用3n表示;依此类推,还有四倍体、五倍体、六倍体……可用4n、5n、6n……表示。凡体细胞中具有二倍以上的染色体数的生物均称为多倍体,包括三倍体、四倍体、五倍体……等”。 “小麦属的多倍体系有一粒小麦,体细胞染色体数2n=14;二粒小麦,体细胞染色体数2n=28,为异源四倍体;普通小麦,体细胞染色体数2n=42,为异源六倍体”。 这种说法并不少见。开始“以n代表基本染色体     

结球甘蓝二、四倍体间杂交三倍体的获得及细胞学鉴定

在对结球甘蓝4x×2x和2x×4x的授粉受精及胚胎发育观察的基础上,结合幼胚离体培养技术,成功地获得了结球甘蓝三倍体材料,并对其减数分裂行为、染色体在后期Ⅰ的分离及雌雄配子的传递率进行了观察研究,这为创建结球甘蓝"初级三体系"等遗传研究奠定了基础.     

枣胚乳三倍体的细胞学－中国果树资源细胞学研究之六

枣属植物经过染色体鉴定的5个种,未发现奇数倍性;中国枣为二倍体,未见自然多倍体类型。1978年通过胚乳培养,已首次诱导出三倍体植株。1983年三倍体始花结果,其细胞学特点如下: 1.胚乳二倍体胚乳二倍体的染色体数2n=24。小孢子母细胞减数分裂染色体行为正常。前期Ⅰ和中期Ⅰ,形成12个二阶体。减数分裂结束,形成正常的四分体,小孢子大小整齐。花粉粒属于正常的三孔沟型。2.胚乳三倍体胚乳三倍体的染色体数2n=36。小孢子母细胞较二倍体大。减数分裂染色体行为不正常。前期Ⅰ和中期Ⅰ有数目不等的单价体、二价体和多价体,染色体群数变动于14—20之间。后期Ⅰ、Ⅱ染色体分离不规则,数目不均衡,有落后染色体,多极分裂,并带有微核。减数分裂结束时,部分小孢子母细胞形成一分体、二分体、不等四分体、五分体和六分体等,小孢子大小不一致。正常花粉比二倍体大,有三孔沟和四孔沟两种类型,还有部分小花粉粒和败育花粉。     

Comparison of methods for hatchery-scale triploidization of European catfish (Silurus glanis L.)

Heat-, cold- and hydrostatic pressure shocks were applied in order to improve triploidy induction in European catfish ( Silurus glanis L . ). A 41°C heat shock (45 s, starting 9 min after gamete activation) provided 88% triploids and a high percentage of malformation (38.8 ± 4.1%). The superior 6°C cold shock (20 min, starting 9 min after gamete activation) gave 100% triploids and a 33.4 ± 3.8% triploid yield. The earliest hydrostatic treatments (600 kg cm²), lasting 4 min and starting 3 min after gamete activation, gave 97.8 ± 1.8% triploids and a 33.7 ± 16.9% triploid yield. The ploidy level was investigated using four approaches: karyotyping, quantification of Ag-stained nucleoli per cell, flow cytometry, and erythrocyte nuclear sizing by computer-assisted image analysis. Induction of triploidy under mass conditions in three experiments gave triploid percentages of 74%, 83% and 66%. Five months later, the percentage of triploids significantly decreased to 12.4%, 8.2% and 21.4%. The growth performance of yearlings was better in diploids than in triploids. Differences between diploids and triploids were 13.5% (NS), 27.6% (P   < 0.001) and 25.4% (P   < 0.05) in the three experiments. Analysis of variance showed the influence of ploidy (P   < 0.001) on growth rate, and multiple range analysis (LSD) assessed differences between total diploids (12.6 g) and total triploids (9.5 g) at the P   < 0.01 level.     

Microsatellite and flow cytometry analysis to help understand the origin of Dioscorea alata polyploids

Background and Aims Dioscorea alata

is a polyploid species with a ploidy level ranging from diploid (2n = 2x = 40) to tetraploid (2n = 4x = 80). Ploidy increase is correlated with better agronomic performance. The lack of knowledge about the origin of D. alata spontaneous polyploids (triploids and tetraploids) limits the efficiency of polyploid breeding. The objective of the present study was to use flow cytometry and microsatellite markers to understand the origin of D. alata polyploids.

Methods

Different progeny generated by intracytotype crosses (2x × 2x) and intercytotype crosses (2x × 4x and 3x × 2x) were analysed in order to understand endosperm incompatibility phenomena and gamete origins via the heterozygosity rate transmitted to progeny.

Results

This work shows that in a 2x × 2x cross, triploids with viable seeds are obtained only via a phenomenon of diploid female non-gametic reduction. The study of the transmission of heterozygosity made it possible to exclude polyspermy and polyembryony as the mechanisms at the origin of triploids. The fact that no seedlings were obtained by a 3x × 2x cross made it possible to confirm the sterility of triploid females. Flow cytometry analyses carried out on the endosperm of seeds resulting from 2x × 4x crosses revealed endosperm incompatibility phenomena.

Conclusions

The major conclusion is that the polyploids of D. alata would have appeared through the formation of unreduced gametes. The triploid pool would have been built and diversified through the formation of 2n gametes in diploid females as the result of the non-viability of seeds resulting from the formation of 2n sperm and of the non-viability of intercytotype crosses. The tetraploids would have appeared through bilateral sexual polyploidization via the union of two unreduced gametes due to the sterility of triploids.     

Development of polyspermic zygote and possible contribution of polyspermy to polyploid formation in angiosperms

Fertilization is a general feature of eukaryotic uni- and multicellular organisms to restore a diploid genome from female and male gamete haploid genomes. In angiosperms, polyploidization is a common phenomenon, and polyploidy would have played a major role in the long-term diversification and evolutionary success of plants. As for the mechanism of formation of autotetraploid plants, the triploid-bridge pathway, crossing between triploid and diploid plants, is considered as a major pathway. For the emergence of triploid plants, fusion of an unreduced gamete with a reduced gamete is generally accepted. In addition, the possibility of polyspermy has been proposed for maize, wheat and some orchids, although it has been regarded as an uncommon mechanism of triploid formation. One of the reasons why polyspermy is regarded as uncommon is because it is difficult to reproduce the polyspermy situation in zygotes and to analyze the developmental profiles of polyspermic triploid zygotes. Recently, polyspermic rice zygotes were successfully produced by electric fusion of an egg cell with two sperm cells, and their developmental profiles were monitored. Two sperm nuclei and an egg nucleus fused into a zygotic nucleus in the polyspermic zygote, and the triploid zygote divided into a two-celled embryo via mitotic division with a typical bipolar microtubule spindle. The two-celled proembryos further developed and regenerated into triploid plants. These suggest that polyspermic plant zygotes have the potential to form triploid embryos, and that polyspermy in angiosperms might be a pathway for the formation of triploid plants.     

Differentiating diploid and triploid individuals using single nucleotide polymorphisms genotyped by amplicon sequencing

Triploidy can occur naturally or be induced in fish and shellfish during artificial propagation in order to produce sterile individuals. Fisheries managers often stock these sterile triploids as a means of improving angling opportunities without risking unwanted reproduction of the stocked fish. Additionally, the rearing of all‐triploid individuals has been suggested as a means to reduce the possibility of escaped aquaculture fish interbreeding with wild populations. Efficient means of determining if an individual is triploid or diploid are therefore needed both to monitor the efficacy of triploidy‐inducing treatments and, when sampling fish from a body of water that has a mixture of diploids and triploids, to determine the ploidy of a fish prior to further analyses. Currently, ploidy is regularly measured through flow cytometry, but this technique typically utilizes a fresh blood sample. This study presents an alternative, cost‐effective method of determining ploidy by analysing amplicon‐sequencing data for biallelic single‐nucleotide polymorphisms (SNPs). For each sample, heterozygous genotypes are identified and the likelihoods of diploidy and triploidy are calculated based on the read counts for each allele. The accuracy of this method is demonstrated using triploid and diploid brook trout (Salvelinus fontinalis) genotyped with a panel of 234 SNPs and Chinook salmon (Oncorhynchus tshawytscha) genotyped with a panel of 298 SNPs following the GT‐seq methodology of amplicon sequencing.     

Why are triploid zebrafish all male?

Adult triploid zebrafish Danio rerio has previously been reported to be all male. This phenomenon has only been reported in one other gonochoristic fish species, the rosy bitterling Rhodeus ocellatus, despite the fact that triploidy is induced in numerous species. To investigate the mechanism responsible, we first produced triploid zebrafish and observed gonad development. Histological sections of juvenile triploid gonads showed that primary growth oocytes were able to develop in the juvenile ovary, but no cortical alveolus or more advanced oocytes were found. All adult triploids examined were male (n = 160). Male triploids were able to induce oviposition by diploid females during natural spawning trials, but fertilization rates were low (1.0 ± 3.1%) compared with diploid male siblings (67.4 ± 16.6%). The embryos produced by triploid sires were aneuploid with a mean ploidy of 2.4 ± 0.1n, demonstrating that triploid males produce aneuploid spermatozoa. After confirming that adult triploids are all male, we produced an additional batch of triploid zebrafish and exposed them (and a group of diploid siblings) to 100 ng/L estradiol (E2) from 5 to 28 dpf. The E2 treated triploids and nontreated triploids were all male. The nontreated diploids were also all male, but the E2 treated diploids were 89% female. This demonstrates that triploidy acts downstream of estrogen synthesis in the sex differentiation pathway to induce male development. Based on this and the observations of juvenile gonad development in triploids, we suggest that triploidy inhibits development of oocytes past the primary growth stage, and this causes female to male sex reversal.     

Gonad Development Characteristics and Sex Ratio in Triploid Chinese Shrimp (<Emphasis Type="Italic">Fenneropenaeus chinensis</Emphasis>)

This paper details for the first time the gonad development characteristics and sex ratio of triploid shrimp (Fenneropenaeus chinensis). In triploid shrimp the development of gonad is apparently impaired, especially in females. In the ovary of triploids, germ cells mainly remain at oogonia stage during September through December. From January to February of the next year, partial primary oocytes developed in the ovary lobes. Spermatocytes and spermatids could be observed in the testes of triploids, and a few sperm were observed in the vas deferens and spermatophores. The morphology of sperms in triploid shrimp was abnormal. Flow cytometry was used to detect the ploidy of sperm in the vas deferens. The data showed that triploidy could affect the sex ratio in Chinese shrimp. The female-to-male ratio in triploids of about 4:1 will favor triploid shrimp aquaculture.     

二倍体鲫鲤F2产生不同倍性卵子的证据

在检测到鲫鲤F2产生3种不同大小(直径分别为0.13 cm,0.17cm和0.2 cm)类型的卵子基础上,进行了F2(♀)×红鲫(♂)及F2(♀)×四倍体鲫鲤(♂)的交配实验.通过染色体计数和流式细胞仪分析,在F2(♀)×红鲫(♂)后代中获得了四倍体、三倍体、二倍体鱼;在F2(♀)×四倍体鲫鲤(♂)后代中获得了四倍体和三倍体鱼.这两个交配组合后代中出现的不同倍性的鱼类为证明鲫鲤F2能产生三倍体、二倍体和单倍体卵子提供了进一步证据.F2(♀)×红鲫(♂)中雄性四倍体鱼的存在说明在四倍体后代中存在基因型为XXXY的个体.对上述两个交配组合后代的四倍体鱼和三倍体鱼的性腺结构观察表明四倍体鱼是可育的,而三倍体鱼是不育的.作者认为鲫鲤F2能够产生二倍体和三倍体卵子与核内复制机制和生殖细胞的融合有关.     

Aneuploidy and genetic variation in the Arabidopsis thaliana triploid response

Polyploidy, the inheritance of more than two genome copies per cell, has played a major role in the evolution of higher plants. Little is known about the transition from diploidy to polyploidy but in some species, triploids are thought to function as intermediates in this transition. In contrast, in other species triploidy is viewed as a block. We investigated the responses of Arabidopsis thaliana to triploidy. The role of genetic variability was tested by comparing triploids generated from crosses between Col-0, a diploid, and either a natural autotetraploid (Wa-1) or an induced tetraploid of Col-0. In this study, we demonstrate that triploids of A. thaliana are fertile, producing a swarm of different aneuploids. Propagation of the progeny of a triploid for a few generations resulted in diploid and tetraploid cohorts. This demonstrated that, in A. thaliana, triploids can readily form tetraploids and function as bridges between euploid types. Genetic analysis of recombinant inbred lines produced from a triploid identified a locus on chromosome I exhibiting allelic bias in the tetraploid lines but not in the diploid lines. Thus, genetic variation was subject to selection contingent on the final ploidy and possibly acting during the protracted aneuploid phase.     

Comparison of genetic variability and parentage in different ploidy classes of the Japanese oyster Crassostrea gigas

Chemical treatments with cytochalasin B were used to induce triploidy in the progeny of a mass fertilization of 3 male and 7 female Crassostrea gigas parents. Triploids were produced either by retention of the first (meiosis I (MI) triploids) or the second (meiosis II (MII) triploids) polar bodies. These animals, together with their diploid siblings, were divided for two experiments. One set was used to compare physiological performance, and the other set deployed to compare growth in two different natural environments. For both experiments, genetic variability in different ploidy classes was estimated using three microsatellite loci and eight allozyme loci. The microsatellite loci were highly polymorphic, allowing independent confirmation of ploidy status and the unambiguous identification of parentage for each oyster. Significant differences in parentage were found between ploidy classes, despite the fact they originated from the same mass fertilization. This indicates that the assumptions of a common genetic background among random samples of animals taken from the same mass fertilization may not be generally valid. Knowledge of parentage also allowed the more accurate scoring of allozyme loci. As expected, triploids were found to be significantly more polymorphic than diploids. However, MI triploids were not significantly more polymorphic than MII triploids. MII triploid genotypes were used to estimate recombination rates between loci and their centromeres. These rates varied between 0.29 and 0.71, indicating only moderate chiasma interference.