Development and application of biological technologies in fish genetic breeding

Fish genetic breeding is a process that remolds heritable traits to obtain neotype and improved varieties.For the purpose of genetic improvement,researchers can select for desirable genetic traits,integrate a suite of traits from different donors,or alter the innate genetic traits of a species.These improved varieties have,in many cases,facilitated the development of the aquaculture industry by lowering costs and increasing both quality and yield.In this review,we present the pertinent literatures and summarize the biological bases and application of selection breeding technologies(containing traditional selective breeding,molecular marker-assisted breeding,genome-wide selective breeding and breeding by controlling single-sex groups),integration breeding technologies(containing cross breeding,nuclear transplantation,germline stem cells and germ cells transplantation,artificial gynogenesis,artificial androgenesis and polyploid breeding)and modification breeding technologies(represented by transgenic breeding)in fish genetic breeding.Additionally,we discuss the progress our laboratory has made in the field of chromosomal ploidy breeding of fish,including distant hybridization,gynogenesis,and androgenesis.Finally,we systematically summarize the research status and known problems associated with each technology.     

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.     

DURATION AND CONSISTENCY OF HISTORICAL SELECTION ARE CORRELATED WITH ADAPTIVE TRAIT EVOLUTION IN THE STREAMSIDE SALAMANDER, AMBYSTOMA BARBOURI

A fundamental challenge in evolutionary biology concerns estimating the extent to which ecological trade-offs may impose constraints on adaptive evolution. Novel ecological stressors may limit adaptive evolution of naive lineages that have experienced historically different selective regimes. Regarded as recently derived from a pond-breeding ancestor, streamside salamanders face the novel and strong selection pressure of breeding in streams with fish predators. A statistical phylogenetic approach was used to test whether adaptive evolution of antipredator performance phenotypes in streamside salamanders was positively associated with: (1) estimated per-lineage duration of coexistence with predatory fish; and (2) consistency of this predator selective-regime within lineages. Average durations of fish contact were computed for each salamander lineage on a set of chronograms. Selection consistency was determined by estimating the number of ecological transitions between fish and fishless states using stochastic character mapping. Historical selection in streamside salamanders can be generally characterized as unstable, apparently punctuated by the stochastic loss and recolonization of predatory fish in most lineages. We found that the efficacy of antipredator phenotypes in salamanders is strongly related to historical duration, as well as consistency, of selection imposed by predatory fish.     

The strange blood: natural hybridization in primates

Hybridization between two closely related species is a natural evolutionary process that results in an admixture of previously isolated gene pools. The exchange of genes between species may accelerate adaptation and lead to the formation of new lineages. Hybridization can be regarded as one important evolutionary mechanism driving speciation processes. Although recent studies have highlighted the taxonomic breadth of natural hybridization in the primate order, information about primate hybridization is still limited compared to that about the hybridization of fish, birds, or other mammals. In primates, hybridization has occurred mainly between subspecies and species, but has also been detected between genera and even in the human lineage. Here we provide an overview of cases of natural hybridization in all major primate radiations. Our review emphasizes a phylogenetic approach. We use the data presented to discuss the impact of hybridization on taxonomy and conservation.     

An Update on Genetic Modification of Chickpea for Increased Yield and Stress Tolerance

Chickpea is a highly nutritious grain legume crop, widely appreciated as a health food, especially in the Indian subcontinent. The major constraints on chickpea production are biotic (Helicoverpa, bruchid, aphid, ascochyta) and abiotic (drought, heat, salt, cold) stresses, which reduce the yield by up to 90%. Various strategies like conventional breeding, molecular breeding, and modern plant breeding have been used to overcome these problems. Conventionally, breeding programs aim at development of varieties that combine maximum number of traits through inter-specific hybridization, wide hybridization, and hybridization involving more than two parents. Breeding is difficult in this crop because of its self-pollinating nature and limited genetic variation. Recent advances in in vitro culture and gene technologies offer unique opportunities to realize the full potential of chickpea production. However, as of date, no transgenic chickpea variety has been approved for cultivation in the world. In this review, we provide an update on the development of genetically modified chickpea plants, including those resistant to Helicoverpa armigera, Callosobruchus maculatus, Aphis craccivora, as well as to drought and salt stress. The genes utilized for development of resistance against pod borer, bruchid, aphid, drought, and salt tolerance, namely, Bt, alpha amylase inhibitor, ASAL, P5CSF129A, and P5CS, respectively, are discussed.     

Rapid formation of distinct hybrid lineages after secondary contact of two fish species (Cottus sp.)

Homoploid hybridization after secondary contact between related species can lead to mixtures of genotypes which have the potential for rapid adaptation to new environmental conditions. Here, we focus on a case where anthropogenic changes within the past 200 years have allowed the hybridization between two fish species (Cottus rhenanus and Cottus perifretum) in the Netherlands. Specifically, we address the question of the dynamics of the emergence of these hybrids and invasion of the river systems. Using a set of 81 mostly ancestry-informative SNP markers, as well as broad sample coverage in and around the area of the initial contact, we find a structured hybrid swarm with at least three distinct hybrid lineages that have emerged out of this secondary contact situation. We show that genetically coherent groups can occur at geographically distant locations, while geographically adjacent groups can be genetically different, indicating that some form of reproductive isolation between the lineages is already effective. Using a newly developed modelling approach, we test the relative influence of founding admixture, drift and migration on the allele compositions of the sampling sites. We find that the allele frequency distributions can best be explained if continued gene flow between the parental species and the hybrid lineages is invoked. Genome mapping of the invasive lineage in the Rhine shows that major chromosomal rearrangements were not involved in creating this distinct lineage. Our results show that hybridization after secondary contact can quickly lead to multiple independent new lineages that have the capacity to form hybrid species.     

Unlocking the barley genome by chromosomal and comparative genomics

We used a novel approach that incorporated chromosome sorting, next-generation sequencing, array hybridization, and systematic exploitation of conserved synteny with model grasses to assign ~86% of the estimated ~32,000 barley (Hordeum vulgare) genes to individual chromosome arms. Using a series of bioinformatically constructed genome zippers that integrate gene indices of rice (Oryza sativa), sorghum (Sorghum bicolor), and Brachypodium distachyon in a conserved synteny model, we were able to assemble 21,766 barley genes in a putative linear order. We show that the barley (H) genome displays a mosaic of structural similarity to hexaploid bread wheat (Triticum aestivum) A, B, and D subgenomes and that orthologous genes in different grasses exhibit signatures of positive selection in different lineages. We present an ordered, information-rich scaffold of the barley genome that provides a valuable and robust framework for the development of novel strategies in cereal breeding.     

Facilitating wide hybridization in Hydrangea s. l. cultivars: A phylogenetic and marker-assisted breeding approach

Hydrangea s. l., belonging to the up-market segment of ornamental cultivars, currently faces a renaissance in horticulture. Hence, novel molecular-assisted breeding approaches are timely. Wide hybridization, i.e. crosses between distantly related species, has been shown to be problematic. Recent studies have considerably improved our knowledge of the phylogenetic relationships between the ornamental Hydrangea s. l. species. A fully resolved and highly supported phylogenetic tree is currently available, based on an extensive marker selection including 13 highly variable chloroplast markers. This robust phylogenetic framework includes the majority of widely cultivated Hydrangea s. l. species that have been the center of attention in a number of crossing projects. The present study is based on this highly supported phylogenetic hypothesis. Here, we aim to select the best candidates for future successful breeding projects, involving interspecific crosses of both closely and distantly related Hydrangea s. l. lineages. Therefore, we integrated the phylogenetic relatedness of potential parental lines along with genetic distances calculated from a wide plastid marker selection. Direct crosses between two species were found to be successful up to an average genetic distance of 0.01065, while failure could be expected at an average genetic distance of 0.01385 and higher. In order to overcome this genetic distance threshold, we propose Hydrangea arborescens, H. sargentiana, H. integrifolia, and H. seemannii as the best candidates for future bridge-cross projects with currently available fertile hybrids. We expect that our results will allow breeders to overcome long-standing wide crossing difficulties and motivate breeding initiatives of potential economic value.     

元基因组文库分析技术研究进展

随着新的分析技术的不断出现和成熟,促进了微生物分子生态学及相关学科的诞生和迅速发展。其中,元基因组文库分析技术即是近年来微生物分子生态学研究领域兴起的一种新的分析技术。就元基因组分析技术诞生的背景及该技术的原理进行了讨论,着重阐述了元基因组文库分析技术在寻找新基因、开发新的生物活性物质、研究群落中微生物多样性、人类元基因组测序等方面的应用。另外,归纳总结了目前国际上常用的诸如PCR为基础的筛选、荧光原位杂交（fluorescent in situ hybridization,FISH）、底物诱导的基因表达筛选（substrate induced gene expression screening,SIGEX）、基因芯片等元基因组文库筛选方法,并就不同方法的优缺点进行了分析和讨论,指出了目前元基因组文库分析技术存在的主要问题并对今后该技术的发展进行了展望。     

基因人工进化的分子育种技术

分子育种技术(molecular breeding technology)利用人工手段模拟生物杂交优势的分子进化模式,在体外构建含有感兴趣基因的大量突变文库,并根据基因表达产物的特性,以合适的筛选方法选择具有最佳功能的改良基因。经过近几年的快速发展,分子育种技术在创造高效随机基因突变库的方法上日臻完善,已在医学、工业和环境保护酶类研究等方面收到了很好的成效。预计未来该技术将广泛地应用于重要传染性疾病(如疟疾、艾滋病病毒等)的预防和治疗,尤其可通过DNA疫苗手段的应用而获得强大的生命力。作者全面地综述了分子育种技术的产生、发展和应用现状。     

鱼类肌间骨发育分子调控机制及遗传选育研究进展

鱼类的肌间骨(Intermuscular bones, IBs),又称肌间小骨,是分布于鱼体两侧肌隔中及连接在椎体上的小骨,仅存在于低等真骨鱼类中。随着鱼类的系统演化,肌间骨的数目和形态也发生着由无到多再到无、由简单到复杂再到简单的变化。中国的大宗淡水鱼类都存在一定数目的肌间骨,严重影响了其食用和加工价值。鉴于鱼类肌间骨重要的系统进化科学研究价值及产业意义,近年来国内外水产研究者围绕鱼类肌间骨分子调控机制和遗传育种开展了系列研究,文章对此进行了系统的阐述,并对其发展前景进行了分析,可为肌间骨发生发育分子机制及少/无肌间骨鱼类新品种的遗传选育研究提供基础资料。     

杂种优势的形成机制及其在动物遗传育种中的应用

在现代化畜禽养殖业中,杂种优势的利用扮演着十分重要的角色。"显性学说""超显性学说"以及"上位性假说"是经典的数量遗传学对杂种优势解释的3种学说。随着遗传学研究和扩展,研究者们发现杂种优势是来自父、母本基因组等位基因之间的相互作用,改变了有关基因的调控网络,才提高了杂交后代的生产繁殖性能。在动物的杂交育种中,一般采用大量品系、品种进行多次试验,进而筛选出所需的优势性状个体,所以适合推广的杂交优势组合难以获得,所以对杂交育种中的不同组合的杂交优势进行准确预测变得格外的重要。     

Biotechnological Approaches for Developing Salt Tolerant Field Crops

Plant cell cultures have been widely used to screen against salt, salt mixtures, sea water and proline analogues, to develop salt tolerant genotypes in crop species. In some cases, selected trait expressed in regenerated plants has also been shown to be inherited through the sexual cycle. Besides, salt-tolerance character available in the wild germ plasm could be introgressed into the economic target species through improved wide hybridization techniques (e.g. embryo rescue, protoplast fusion, etc.). The third approach makes use of the recombinant DNA technology for identification, cloning, transfer and expression of the salt-tolerance genes in the target species. Basic research on salt-tolerance mechanisms at molecular level, stress related proteins and DNA sequences, plant morphogenesis, somatic cell hybridization, developmental regulation and selection strategies, however, need to be intensified to accelerate breeding for this complex characteristic.     

Genome Editing—Principles and Applications for Functional Genomics Research and Crop Improvement

Genome editing technologies are powerful tools for studying gene function and for crop improvement. The technologies rely on engineered endonucleases to generate double stranded breaks (DSBs) at target loci. The DSBs are repaired through the error-prone non-homologous end joining (NHEJ) and homology-directed repair (HDR) pathways in cells, resulting in mutations and sequence replacement, respectively. In the widely used CRISPR/Cas9 system, the endonuclease Cas9 is targeted by a CRISPR small RNA to DNA sequence of interest. In this review, we describe the four available types of genome editing tools, ZFN, TALEN, CRISPR/Cas9 and CRISPR/Cpf1, and show their applications in functional genomics research and precision molecular breeding of crops.     

Historical introgression among the American live oaks and the comparative nature of tests for introgression

Introgressive hybridization challenges the concepts we use to define species and infer phylogenetic relationships. Methods for inferring historical introgression from the genomes of extant species, such as ABBA‐BABA tests, are widely used, however, their results can be easily misinterpreted. Because these tests are inherently comparative, they are sensitive to the effects of missing data (unsampled species) and nonindependence (hierarchical relationships among species). We demonstrate this using genomic RADseq data sampled from all extant species in the American live oaks (Quercus series Virentes), a group notorious for hybridization. By considering all species and their phylogenetic relationships, we were able to distinguish true hybridizing lineages from those that falsely appear admixed. Six of seven species show evidence of admixture, often with multiple other species, but which is explained by introgression among a few related lineages occurring in close proximity. We identify the Cuban oak as the most admixed lineage and test alternative scenarios for its origin. The live oaks form a continuous ring‐like distribution around the Gulf of Mexico, connected in Cuba, across which they could effectively exchange alleles. However, introgression appears highly localized, suggesting that oak species boundaries and their geographic ranges have remained relatively stable over evolutionary time.     

Chromosome changes after polyploidization in Triticeae

Chromosome changes are common in Triticeae, and they occur widely in natural and resynthesized polyploidy. Two important factors, nucleocytoplasmic interaction (internal) and the environment (external), can significantly influence chromosome changes after polyploidization. And chromosomal DNA changes play key roles during the initial formation, gradual stabilization, and establishment of polyploids. Hybrid breeding between common wheat and related wild species of Triticeae is an example of polyploidization, and many of the chromosome changes occurring after hybridization could be useful for improving wheat varieties. The famous chromosomal translocation 1BL/1RS that occurred after ancestral hybridization between wheat and rye is distributed widely among modern wheat varieties and makes a big contribution to wheat breeding; xiaoyan 6 is a similarly distant hybridization between wheat and Agropyron elongatum (Host) P. Beauv. in China. This chromosome translocation line was cultivated as the main variety in Shaanxi Province for 16 years and has also been used as a core parent for wheat breeding in China during the past 20 years because of its outstanding merits.     

微卫星标记及其在蚜虫种群生物学研究中的应用

微卫星（SSR）是以1～6个碱基为重复单位组成的简单串联重复序列,具有丰度高、多态性高、共显性遗传、选择中性和可重复性好等优点。微卫星标记技术可分为三种类型：基于杂交的SSR指纹,基于PCR的SSR指纹和单位点SSR的PCR扩增。前两者不需要知道基因组序列信息,属于多位点标记;后者则需要前期工作,属于单位点标记,即通常所指的微卫星标记。到目前为止,共在14种蚜中筛选到141个微卫星位点,并发表了相关的引物序列,这给今后的相关研究提供了丰富的共享资源。业已证明微卫星引物在邻近种之间有一定的通用性。微卫星标记严格遵守孟德尔遗传规律,已经被用来推断某些蚜虫有性生殖的情况。同种蚜虫中可能同时存在有性系和无性系,微卫星研究证明有性系比无性系有更高的遗传多样性,无性系通常表现出杂合过剩和连锁不平衡。蚜虫的迁飞规律适合用微卫星标记加以研究,已有的研究显示在具有高度迁飞特性的种类中,地理种群间的遗传分化程度低,基因频率的相似性高。存在广泛分布的相同的多位点基因型是迁飞性蚜虫的另一个重要特征。在我国,微卫星应用于蚜虫生物学研究的工作还较少,鉴于该种标记的优良特性和巨大的潜力,本文建议今后相关的研究应该首先考虑微卫星标记。     

Expansion of the genomics research on Atlantic salmon Salmo salar L. project (GRASP) microarray tools

Salmonids are the most widely studied group of fish, and in the last few years, genomics technologies have begun to contribute to this rich biology. The first salmonid microarrays appeared in 2004 and since then several dozen studies have demonstrated the utility of genomic approaches. The widespread use of the genomics research on Atlantic salmon project 16 k array and greatly expanded genome resources have led to the development of an experimental 5 k oligo (70-mer) array and a 32 k cDNA microarray in the near future. In this paper, the authors examined some of the procedures used in the development of past arrays and reexamined them in light of new genomic data available. Some preliminary control experiments of the new 5 k array were investigated that examine oligo designs based on distance from the polyA tail, the effects of mismatches and cross-species hybridization specificity. Beneficial approaches are also identified in the development of the new 32 k cDNA array.