后基因组时代基因组重排在微生物菌种选育中的应用及展望

基因组重排作为一种实用高效的育种技术,在缺乏遗传背景认知和可操作遗传体系等条件下,可以突破微生物种属间的限制,经过多轮递推的原生质体融合来加速其人工定向进化,在微生物菌种改良及代谢产物开发和产业化等研究领域得到了广泛应用。步入后基因组时代,快速发展的组学和生物信息学使基因组重排成为连接各种微生物育种方法的重要纽带,为我们深入探索微生物复杂的代谢网络和全局调控机制,更为精准地实施对微生物的人工调控和定向进化提供了契机。本文系统性地回顾了近年来基因组重排在微生物菌种选育中的应用研究,尤其针对围绕其开展的组学研究进行了详细阐述,并对基因组重排与组学、生物信息学和合成生物学等新兴技术的联合应用进行了展望。     

基因组选择及其应用

品种选育在农业生产中占有十分重要的地位,育种值估计是品种选育的核心。随着遗传标记的发展,尤其是高通量的基因分型技术,使得从基因组水平估计育种值成为可能,即基因组选择。文章将基因组选择的方法分为两类:一是基于估计等位基因效应来预测基因组估计育种值(GEBV),如最小二乘法,随机回归-最佳线性无偏预测(RR-BLUP)、Bayes、主成分分析等方法;二是基于遗传关系矩阵来预测GEBV,通过采用高通量标记构建个体间的遗传关系矩阵,然后用线性混合模型来预测育种值,即GBLUP法,并以这两种分类简要介绍了基因组选择各种方法的大致原理。影响基因组选择准确性的因素主要有标记类型和密度、单倍型长度、参考群体大小和标记-数量性状基因座(QTL)连锁不平衡(LD)大小等;在基因组选择的各种方法中,一般说来Bayes方法和GBLUP方法具有较高的准确性,最小二乘法最差;GBLUP计算速度快,能够将标记和系谱结合起来,因而比其他方法更具优势。尽管基因组选择取得了很大进展,但在理论方面还面临着一些挑战,如联合育种、长期选择的遗传进展及如何解析与性状有关和无关的标记等。基因组选择在一些动植物育种上已经开始应用,在人类遗传倾向预测和进化动力学研究中也有潜在的应用前景。基因组选择在个体间亲缘关系的量化上有了突破,比传统方法更加精确,因此,基因组选择将会是动植物育种史上革命性的事件。     

BREEDING RESOURCE DISTRIBUTION AFFECTS SELECTION GRADIENTS ON MALE PHENOTYPIC TRAITS: EXPERIMENTAL STUDY ON LIFETIME REPRODUCTIVE SUCCESS IN THE BITTERLING FISH (RHODEUS AMARUS)

The spatial distribution of breeding resources can have pronounced demographic and evolutionary consequences. We used 20 experimental groups of the bitterling ( Rhodeus amarus ), an annual fish with a promiscuous, resource-based mating system, and extended breeding season to investigate how the spatial distribution (clumped or regular) of bitterling oviposition sites (live freshwater mussels) affected offspring production, variation in reproductive success, and directional selection on phenotypic traits over their entire reproductive lifetime. We did not detect any effect of resource distribution on offspring production or variation in reproductive success among individual fish, although variation between replicates was higher with a clumped distribution. This finding is discussed with regard to the incidence of alternative mating behaviors (sneaking) within the limitations imposed by our experimental design. Breeding resource distribution had a significant effect on selection on male phenotypic traits. Stronger directional selection on traits associated with intrasexual competition for fertilizations, gonad mass (an indicator of sperm competition), and the extent of red, carotenoid-based pigment in the iris (an index of dominance status), was detected with a clumped resource distribution. With a regular resource distribution, a stronger positive selection on male body size was detected. We discuss the implications of our results for natural populations.     

Biotechnology: Genetic improvement of sorghum (Sorghum bicolor (L.) Moench)

Summary This report reviews the contributions to the improvement of sorghum (Sorghum bicolor (L.) Moench) through traditional approaches with emphasis on the application of biotechnological methods. Strategies include breeding for higher yield, improved grain quality, and biotic and abiotic stress tolerance. Hybrid development and polyploidy breeding are also discussed. Plant breeders, working in concert with biotechnologists, have developed new powerful tools for plant genetic manipulation and genotype evaluation that will significantly improve the efficiency of plant breeding. Improving sorghum through biotechnology is the latest in a long series of technologies that have been applied to this crop. Five basic tools of technology have been developed for sorghum improvement: (1) in vitro protocols for efficient plant regeneration; (2) molecular markers; (3) gene identification and cloning; (4) genetic engineering and gene transfer technology to integrate desirable traits into the sorghum genome; and (5) genomics and germplasm databases. Reports on studies involving the problems, progress, and prospects for utilizing the biotechnological methods for sorghum improvement are discussed.     

Microglia of teleosts: facing a challenge in neurobiology

This review is concerned with recent literature on teleost fish CNS microglia. It covers not only various aspects of these cells, notably comparing them with mammalian microglia, but also points out the several potentialities neural tissue of teleosts exhibits in neurobiological research. The relationships between neurons and glial cells are considered in fish, aiming at an integrated picture of the complex ways neurons and glia communicate and collaborate in normal and injured neural tissues. In addition, attention has been paid to different teleost models according to their availability, easy maintenance in experimental conditions, possibilities of embryos manipulation and sequenced genome. The recent setting up of successful protocols for fish glia and mixed neuron-glia cultures, together with the molecular facilities offered from genome knowledge, should provide a new boost to studies about microglia and neuron-microglia relationships.     

Pair-bond formation and breeding-site limitation in the convict cichlid, <Emphasis Type="Italic">Archocentrus nigrofasciatus</Emphasis>

Convict cichlids (Archocentrus nigrofasciatus) form pair bonds, are serially monogamous, and give biparental care to their offspring. Males and females may individually establish a breeding site before courtship, or they may form a pair and then obtain a breeding site together. Such sites are important resources for convict cichlids, and their availability often limits reproduction. Here, we examined if variation in breeding-site availability affected pair bonding between males and females and aggressive behaviors between bonded pairs. Groups of three males and three females were placed in large aquaria and randomly assigned to the following treatments: (1) the fish were provided with two breeding sites; (2) the fish were provided with two breeding sites throughout the experiment, one of which was later removed; (3) the fish were provided with one breeding site initially, and a second was later added; (4) the fish were provided with no breeding sites throughout the experiment. We found that pair bonds rarely formed when no breeding sites were provided; however, pair-bond formation was not limited by breeding-site number when at least one breeding site was available. In addition, there were no differences in aggression by paired individuals after breeding site manipulation, perhaps indicating that changes in the subsequent availability of breeding sites after pair formation did not influence a pair’s aggression.     

Allozyme polymorphisms in tetraploid potato gene pools and the effect on human selection

The need for broadening a crop’s genetic base may be determined by comparing allele frequencies within the gene pools of farmer selections in their centers of diversity with that of modern breeding populations. The genetic structure of Andean and Chilean potato farmer selections was investigated with the aid of nine isozymes, which have been studied in detail and used to characterize North American cultivars and advanced breeding lines. These isozymes are associated with the most-important agronomic or quality characters in the North American gene pool. By comparing these data with previous analyses of the North American gene pool, allozyme frequency changes for nine loci were monitored. Allozyme frequency changes were not always due to genetic drift, but resulted also from directional selection of isozyme marker linked quantitative trait loci (QTLs) affecting agronomic or quality characters. Changes in allozyme frequency can also occur as a consequence of pleiotropy, i.e. the isozyme itself may be involved in the expression of a phenotype. These allozyme frequency changes may reflect the manipulation of the potato genome by breeders. There were allozymes in some North American cultivars that were not observed in the farmer selections from the Andes and Chile. This confirms that breeders have already introgressed exotic genes from wild and other primitive cultivated tuber-bearing Solanum species. On this basis, the need for broadening the genetic base for specific chromosomes (or chromosome regions) should be based on analysis with these and other genetic markers available in potato. Received: 20 November 2000 / Accepted: 27 December 2000     

Establishment and application of distant hybridization technology in fish

Hybridization is widely used. However, for a long time, systematic theories and technologies related to hybridization in fish have been lacking. In this study, through long-term systematic research, we investigated and obtained the main rules regarding inheritance and reproduction related to fish distant hybridization. Furthermore, we established one-step and multistep breeding technologies that were suitable for interspecific hybridization and intraspecific hybridization. Simultaneously, we used these two breeding technologies to produce a batch of diploid fish lineages and tetraploid fish lineages and improved fishes. In addition, we widely discuss the methods, technologies and results of hybridization breeding, referring to the domestic and foreign literature on fish hybridization. We hope that this paper will be beneficial for the research and application of fish hybrid breeding.     

Whole-genome strategies for marker-assisted plant breeding

Molecular breeding for complex traits in crop plants requires understanding and manipulation of many factors influencing plant growth, development and responses to an array of biotic and abiotic stresses. Molecular marker-assisted breeding procedures can be facilitated and revolutionized through whole-genome strategies, which utilize full genome sequencing and genome-wide molecular markers to effectively address various genomic and environmental factors through a representative or complete set of genetic resources and breeding materials. These strategies are now increasingly based on understanding of specific genomic regions, genes/alleles, haplotypes, linkage disequilibrium (LD) block(s), gene networks and their contribution to specific phenotypes. Large-scale and high-density genotyping and genome-wide selection are two important components of these strategies. As components of whole-genome strategies, molecular breeding platforms and methodologies should be backed up by high throughput and precision phenotyping and e-typing (environmental assay) with strong support systems such as breeding informatics and decision support tools. Some basic strategies are discussed in this article, including (1) seed DNA-based genotyping for simplifying marker-assisted selection (MAS), reducing breeding cost and increasing scale and efficiency, (2) selective genotyping and phenotyping, combined with pooled DNA analysis, for capturing the most important contributing factors, (3) flexible genotyping systems, such as genotyping by sequencing and arraying, refined for different selection methods including MAS, marker-assisted recurrent selection and genomic selection (GS), (4) marker-trait association analysis using joint linkage and LD mapping, and (5) sequence-based strategies for marker development, allele mining, gene discovery and molecular breeding.     

Genome editing of polyploid crops: prospects,achievements and bottlenecks

Plant breeding aims to develop improved crop varieties. Many crops have a polyploid and often highly heterozygous genome, which may make breeding of polyploid crops a real challenge. The efficiency of traditional breeding based on crossing and selection has been improved by using marker-assisted selection (MAS), and MAS is also being applied in polyploid crops, which helps e.g. for introgression breeding. However, methods such as random mutation breeding are difficult to apply in polyploid crops because there are multiple homoeologous copies (alleles) of each gene. Genome editing technology has revolutionized mutagenesis as it enables precisely selecting targets. The genome editing tool CRISPR/Cas is especially valuable for targeted mutagenesis in polyploids, as all alleles and/or copies of a gene can be targeted at once. Even multiple genes, each with multiple alleles, may be targeted simultaneously. In addition to targeted mutagenesis, targeted replacement of undesirable alleles by desired ones may become a promising application of genome editing for the improvement of polyploid crops, in the near future. Several examples of the application of genome editing for targeted mutagenesis are described here for a range of polyploid crops, and achievements and bottlenecks are highlighted.

     

Genome sequence of Corynebacterium glutamicum ATCC 14067, which provides insight into amino acid biosynthesis in coryneform bacteria

We report the genome sequence of Corynebacterium glutamicum ATCC 14067 (once named Brevibacterium flavum), which is useful for taxonomy research and further molecular breeding in amino acid production. Preliminary comparison with those of the reported coryneform strains revealed some notable differences that might be related to the difficulties in molecular manipulation.     

我国淡水养殖鱼类育种的实践和思考

用于鱼类育种的传统方法 ,如引种驯化、选种和杂交 ,虽然有其自身的局限性 ,但都是行之有效的 .自本世纪 70年代以来兴起的生物工程技术 ,已应用于鱼类育种 ,如细胞工程中的雌核发育、多倍体形成和性别控制等方面都取得了较好的成效 .80年代以来 ,基因工程技术也已用之于鱼类育种的研究中 ,并取得了一些成效 .实践证明 ,基因工程育种离生产尚有一定距离 .根据当前的实际 ,鱼类育种仍然离不开传统方法 ,应该是细胞工程和传统方法相结合 ,或是细胞工程和基因工程相结合 ,或是传统方法、细胞工程和基因工程相结合的综合技术育种     

Prospects for application of breakthrough technologies in breeding: The CRISPR/Cas9 system for plant genome editing

Integration of the methods of contemporary genetics and biotechnology into the breeding process is assessed, and the potential role and efficacy of genome editing as a novel approach is discussed. Use of molecular (DNA) markers for breeding was proposed more than 30 years ago. Nowadays, they are widely used as an accessory tool in order to select plants by mono- and olygogenic traits. Presently, the genomic approaches are actively introduced into the breeding processes owing to automatization of DNA polymorphism analyses and development of comparatively cheap methods of DNA sequencing. These approaches provide effective selection by complex quantitative traits, and are based on the full-genome genotyping of the breeding material. Moreover, biotechnological tools, such as doubled haploids production, which provides fast obtainment of homozygotes, are widely used in plant breeding. Use of genomic and biotechnological approaches makes the development of varieties less time consuming. It also decreases the cultivated areas and financial expenditures required for accomplishment of the breeding process. However, the capacities of modern breeding are not limited to only these advantages. Experiments carried out on plants about 10 years ago provided the first data on genome editing. In the last two years, we have observed a sharp increase in the number of publications that report about successful experiments aimed at plant genome editing owing to the use of the relatively simple and convenient CRISPR/Cas9 system. The goal of some of these experiments was to modify agriculturally valuable genes of cultivated plants, such as potato, cabbage, tomato, maize, rice, wheat, barley, soybean and sorghum. These studies show that it is possible to obtain nontransgenic plants carrying stably inherited, specifically determined mutations using the CRISPR/Cas9 system. This possibility offers the challenge to obtain varieties with predetermined mono- and olygogenic traits.     

Advances in molecular methods to alter chromosomes and genome in the yeast <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

A fundamental issue in biotechnology is how to breed useful strains of microorganisms for efficient production of valuable biomaterials. On-going and more recent developments in gene manipulation technologies and chromosomal and genomic modifications in particular have facilitated important contributions in this area. “Chromosome manipulation technology” as an outgrowth of “gene manipulation technology” may provide opportunities for creating novel strains of organisms with a variety of genomic constitutions. A simple and rapid chromosome splitting technology called “PCR-mediated chromosome splitting” (PCS) that we recently developed has made it possible to manipulate chromosomes and genomes on a large scale in an industrially important microorganism, Saccharomyces cerevisiae. This paper focuses on recent advances in molecular methods for altering chromosomes and genome in S. cerevisiae featuring chromosome splitting technology. These advances in introducing large-scale genomic modifications are expected to accelerate the breeding of novel strains for biotechnological purposes, and to reveal functions of presently uncharacterized chromosomal regions in S. cerevisiae and other organisms.     

Pedigree‐based analysis of derivation of genome segments of an elite rice reveals key regions during its breeding

Analyses of genome variations with high‐throughput assays have improved our understanding of genetic basis of crop domestication and identified the selected genome regions, but little is known about that of modern breeding, which has limited the usefulness of massive elite cultivars in further breeding. Here we deploy pedigree‐based analysis of an elite rice, Huanghuazhan, to exploit key genome regions during its breeding. The cultivars in the pedigree were resequenced with 7.6× depth on average, and 2.1 million high‐quality single nucleotide polymorphisms (SNPs) were obtained. Tracing the derivation of genome blocks with pedigree and information on SNPs revealed the chromosomal recombination during breeding, which showed that 26.22% of Huanghuazhan genome are strictly conserved key regions. These major effect regions were further supported by a QTL mapping of 260 recombinant inbred lines derived from the cross of Huanghuazhan and a very dissimilar cultivar, Shuanggui 36, and by the genome profile of eight cultivars and 36 elite lines derived from Huanghuazhan. Hitting these regions with the cloned genes revealed they include numbers of key genes, which were then applied to demonstrate how Huanghuazhan were bred after 30 years of effort and to dissect the deficiency of artificial selection. We concluded the regions are helpful to the further breeding based on this pedigree and performing breeding by design. Our study provides genetic dissection of modern rice breeding and sheds new light on how to perform genomewide breeding by design.     

An experiment of fish spillover from a marine reserve in Cuba

Several studies on adult fish movement from marine protected areas to zones open to fishing activity conclude spillover is present, but most of these investigations use indirect evidence and small-sized species of little commercial importance. This paper reports the effects of manipulating a density gradient on movements of large-sized and commercially-important fish across “Jardines de la Reina” Marine Reserve boundaries, using tagging methods and visual census. Tagging was carried out using dart tags and modified spearguns at an experimental and a control site. Density of fish was experimentally manipulated on the unprotected side of the boundary. Before experimental manipulation, fish density was similar in both experimental and control sites and on both sides of the boundaries. After manipulation, fish density in the unprotected side of experimental site declined dramatically and a strong gradient was established through the boundary. One month later, this forced gradient disappeared, returning to the situation at the beginning of the study. This last result is due to spillover effect: the mean distance traveled by fish increased 1.5 times (mean from below 200 m to more than 300 m), the mean emigration rate doubled and the immigration rate decreased, allowing density levels to recover after manipulation.     

鱼类雄核发育的研究进展

雄核发育是在雄核控制下发育的一种特殊的有性生殖方式,它是鱼类单性发育的重要组成部分。本文全面综述了国内外鱼类雄核发育研究的进展状况。主要包括雌核染色体灭活和雄核二倍化诱导技术。雌核染色体灭活主要通过γ、X和紫外线(UV)等辐射处理完成的,其辐射剂量和方法因研究者不同而不同。雄核的二倍化诱导则主要是采用静水压和温度休克阻止第一次有丝分裂来实现的,其二倍化诱导条件与研究的种类不同而有所差异。同时,本文对鱼类雄核发育在遗传育种中的应用前景作了评价。 Abstract：Fish andtogenesis is a kind of secial sexual development which is controlled by male nucleus and it is an important part of fish unisexual development. This paper summarizes the progress of fish androgenesis comprehensively in the world.It includes the techniques of inactive female nucleus of eggs and diploidization of male nucleus. The methods of inactive female nucleus are carride out by using γ,X and ultraviolet (UV)rays to irradiate the eggs,the irradiation dosages and methods are different based on the different researchers. The techniques of diploidization of male nucleus are carried out by using hydrostatic pressure, heat and cold shock to restrain the fish mitosis,the diploidization condition are different from species to species.Meanwhile,the paper evaluates the application perspectives of fish androgenesis on the genetics and breeding.     

Near‐complete genome assembly and annotation of the yellow drum (Nibea albiflora) provide insights into population and evolutionary characteristics of this species

Yellow drum (Nibea albiflora) is an important fish species in capture fishery and aquaculture in East Asia. We herein report the first and near‐complete genome assembly of an ultra‐homologous gynogenic female yellow drum using Illumina short sequencing reads. In summary, a total of 154.2 Gb of raw reads were generated via whole‐genome sequencing and were assembled to 565.3 Mb genome with a contig N₅₀ size of 50.3 kb and scaffold N₅₀ size of 2.2 Mb (BUSCO completeness of 97.7%), accounting for 97.3%–98.6% of the estimated genome size of this fish. We further identified 22,448 genes using combined methods of ab initio prediction, RNAseq annotation, and protein homology searching, of which 21,614 (96.3%) were functionally annotated in NCBI nr, trEMBL, SwissProt, and KOG databases. We also investigated the nucleotide diversity (around 1/390) of aquacultured individuals and found the genetic diversity of the aquacultured population decreased due to inbreeding. Evolutionary analyses illustrated significantly expanded and extracted gene families, such as myosin and sodium: neurotransmitter symporter (SNF), could help explain swimming motility of yellow drum. The presented genome will be an important resource for future studies on population genetics, conservation, understanding of evolutionary history and genetic breeding of the yellow drum and other Nibea species.     

Research advances in animal distant hybridization

Distant hybridization refers to crosses between two different species or higher-ranking taxa that enables interspecific genome transfer and leads to changes in phenotypes and genotypes of the resulting progeny. If progeny derived from distant hybridization are bisexual and fertile, they can form a hybrid lineage through self-mating, with major implications for evolutionary biology, genetics, and breeding. Here, we review and summarize the published literature, and present our results on fish distant hybridization. Relevant problems involving distant hybridization between orders, families, subfamilies, genera, and species of animals are introduced and discussed, with an additional focus on fish distant hybrid lineages, genetic variation, patterns, and applications. Our review serves as a useful reference for evolutionary biology research and animal genetic breeding.