食用菌原生质体融合育种研究进展

原生质体融合是微生物遗传育种中的一项重要技术,可以在种间、属间及科间构建出新型菌株,同时也是菌种改良的重要手段之一,在遗传育种中具有广阔的应用前景.从食用菌原生质体融合技术在育种上的特点,原生质体制备及影响因素,融合方法,融合子的鉴定以及前景展望等方面进行了综述.     

酿酒酵母原生质体的融合

原生质体融合技术开始于动植物细胞,特别是Willin等报道了PEG有助于高等植物细胞原生质体融合以后,这一技术广泛地应用于植物、微生物原生质体的融合和动物细胞的融合。原生质体的制备、培养、融合和发育为研究细胞分化、膜结构与功能、定向育种等生物学     

原生质体技术在工业微生物育种中的应用

原生质体技术已成为工业微生物育种手段之一。本文着重于工业微生物原生质体制备,促融因子及重组体选择策略等进行较详细的讨论,最后提出了若干建议。     

棒状杆菌灭活原生质体种间融合

原生质体融合技术越来越广泛地应用于遗传学基础理论和工业育种研究,且融合的方法不断改进。近来报道用灭活的亲株原生质体融合也能达到遗传重组:用链霉素灭活枯草芽孢杆菌原生质体、热灭活巨大芽孢杆菌原生     

原生质体融合技术及在微生物育种中的应用

原生质体融合又称细胞融合,具有遗传信息传递量大,不受亲缘关系的影响,可以集双亲优良遗传性状为一体,并定向筛选表现双亲遗传性状的融合子,操作简便等优点,具有广阔的应用前景。本文就原生质体融合的程序以及该技术在微生物育种中的应用作一综述。     

地衣芽孢杆菌原生质体最佳形成和再生条件的研究

近年来,微生物原生质体融合及诱变受到广泛重视,实验方法日趋完善,实践证明它是一种有效的育种方法。在微生物原生质体制备和再生过程中,各种微生物所需的最佳条件差别很大。我们研究厂地衣芽孢杆菌53-A,菌株原生质体形成和再生的各种因素及最佳条件,并用光学显微镜和电镜观察了原生质体形成、再生和原生质体的聚集现象,现将结果报道如下。     

基因组改组技术及其在微生物育种中的应用研究进展

Genome shuffling(基因组改组)技术是借助递归式原生质体融合策略对微生物基因组进行遗传改良的一种新兴微生物育种方法。自2002年首次被用来培育tylosin高产菌株以来,目前已为育种工作者广泛采用。对Genome shuffling技术的原理及最近的应用研究进展进行了综述,探讨了其局限性,并展望了其发展的趋势。     

大型真菌原生质体融合技术研究进展

原生质体融合技术是近些年来迅速发展起来的细胞工程育种新技术,它具有许多常规杂交方法所无法比拟的独到之处。随着生物学研究手段的不断创新,使得原生质体融合技术正在逐步完善。本文综述了原生质体融合技术在大型真菌遗传方面的研究进展。     

原生质体融合技术在香菇育种中的应用

对香菇原生质体融合技术以及其融合育种研究作了简要介绍,并初步阐明原生质体融合育种的理论与实际意义。     

念珠菌属原生质体形成与融合的基础研究

一本文对主要条件致病性念珠菌属进行了原生质体形成.再生及种间:属间融合的基础研究。对融合产物进行了生物特性的观察。结果表明:念珠菌属可形成原生质体,其原生质体在一定条件下可互栩融合,其融合产物的生物特性可与亲株不同。白色念珠菌与其它种间,酵母菌属间的融合产物,对小白鼠无致病性。细胞融合是近年来发展起来的一项微生物育种新技术,它是通过两亲株原生质体融合而达到杂交目的的(王俊英,1982)。在细胞工程中,原生质体分离、培养、融合和发育技术是十分重要的。为将这一技术应用于防病治病,我们试对主要条件致病性念珠菌属进行了原生质体的分离.再生及融合的基础研究。     

禾本科三倍体: 形成、鉴定与利用

禾本科三倍体的形成途径包括2n配子融合、倍性间杂交、多精受精和胚乳培养。其中, 2n配子融合和倍性间杂交分别为自然界和人工合成三倍体的主要途径。该文介绍了形态学观测、染色体分析、流式细胞术和分子标记等倍性鉴定方法在禾本科三倍体中的应用及其优缺点。目前, 三倍体在禾谷类作物中无直接应用价值, 但可作为通往多倍体、非整倍体和转移异源基因的遗传桥梁。多年生禾本科三倍体(特别是异源三倍体)在饲草或能源作物中已得到广泛应用, 在该类型禾本科作物中均可直接尝试三倍体育种。多倍体的三倍体育种和无融合生殖三倍体育种可作为未来禾本科三倍体的研究方向。三倍性胚乳培养可以一步合成三倍体, 多精受精可以实现遗传上3个不同基因组的一步融合, 在三倍体研究中应予以重视。鉴于2n配子融合、多精受精的稀有特性和倍性间杂交、胚乳培养频繁的染色体变异, 高通量三倍体鉴定技术的发展将是三倍体研究实现突破的关键。     

Protoplast Fusion in Banana (<Emphasis Type="Italic">Musa</Emphasis> spp.): Comparison of Chemical (PEG: Polyethylene Glycol) and Electrical Procedure

Optimization of protoplast fusion parameters is a prerequisite for the establishment of somatic fusion technology for banana breeding. In the present investigations, we compared the most frequently used fusion methods: the electrofusion technique and chemical procedure (polyethylene glycol). With regard to frequency of binary fusion, protoplast fusion with the fusogen polyethylene glycol was best. Conversely, electric fusion was found to be better with respect to mitotic activities, somatic embryogenesis and plantlet regeneration rate.     

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。     

Protoplast fusion for production of tetraploids and triploids: applications for scion and rootstock breeding in citrus

Protoplast fusion technology has been utilized in many crops to generate allotetraploid somatic hybrids, and sometimes autotetraploids as a byproduct of the process. A brief history of this technology development is provided, along with a simple protocol developed for citrus, which can be easily adapted to other plants. Protoplast fusion has become a significant tool in ploidy manipulation that can be applied in various cultivar improvement schemes. In rare cases, a new somatic hybrid may have direct utility as an improved cultivar; however, the most important application of somatic hybridization is the building of novel germplasm as a source of elite breeding parents for various types of conventional crosses for both scion and rootstock improvement. Somatic hybridization is generating superior allotetraploid breeding parents for use in interploid crosses to generate seedless triploids. Seedlessness is a primary breeding objective for new fresh fruit citrus varieties, and several thousand triploid hybrids have been produced using somatic hybrids as the tetraploid parent. Protoplast fusion is also being utilized to produce somatic hybrids that combine complementary diploid rootstocks, which have shown good potential for tree size control. Tree size control has gained importance as a means of reducing harvesting costs, maximizing the efficiency of modern cold protection methodology, and facilitating the adaptation of new fruit production systems. Successful somatic hybridization in citrus rootstock improvement has enabled rootstock breeding at the tetraploid level via sexual hybridization, which can yield maximum genetic diversity in zygotic progeny upon which to impose selection for the many traits required in improved rootstock cultivars, including disease and insect resistance, broad adaptation, tree size control, and the ability to consistently produce high yields of quality fruit. Recent progress and successful examples of these applications are discussed. Finally, a discussion of the genetic potential of somatic hybrids as breeding parents, including meiotic behavior and inheritance is provided.     

斑玉蕈育种中漆酶转化体系建立的初步研究

漆酶具有降解木质素,氧化降解酚类物质,抑制杂菌,改善出菇品质等作用。采用酶学与原生质体融合相结合的技术,建立斑玉蕈育种中漆酶转化体系,将漆酶活性较高、生长速度较快的凤尾菇原生质体经高温灭活与斑玉蕈原生质体融合,通过RB-PDA平板显色技术筛选出具有漆酶活性较高的两融合菌株Ⅲ18C、Ⅲ2A,并对其进行了拮抗试验、RAPD分子标记、漆酶基因扩增等研究。结果表明,筛选出的两融合菌株与两亲本具有明显的拮抗线,随即引物扩增的条带与两亲本有明显的差异,并扩增出漆酶基因的一个片段。同时也表明利用漆酶转化体系筛选融合菌株具有目标明确、准确、快速的优点。     

Genetic improvement of Citrus for disease resistance

Progress in the genetic improvement of Citrus species was reviewed. Tools used for the genetic improvement of Citrus were categorised as conventional (introduction, selection and hybridisation) and non-conventional methods (mutation, somatic cell hybridisation and genetic engineering) of improvement. Genes linked with the disease resistance were characterised and tagged through molecular marker techniques such as Sequenced Characterised Amplified Region and Cleaved Amplified Polymorphic Sequences. Disease resistance genes showed both monogenic and polygenic inheritance. Conventional methods for disease resistance improvement of Citrus were bottleneck due to inadequate and lengthy breeding procedures. However, non-conventional methods, such as mutation breeding and protoplast fusion, have been routinely utilised for the production of disease resistant germplasm while novel genes from variable sources were used to transform Citrus species to induce resistance against diseases. These non-conventional techniques have been shown to overcome the disadvantages of conventional breeding procedures and could be regarded as rapid methods of genetic improvement as well as helpful to overcome the interspecies barrier.     

枯草芽胞杆菌基因组混组方法

基因组混组作为一种育种方法,通过循环原生质体融合等手段,使得不同菌株来源的基因组能够得到充分重组,增加将正向突变整合到同一重组子中的机会。使用4株带有4种不同标记的枯草芽胞杆菌亲本为初始菌株,通过循环转化、循环转导或循环原生质体融合的手段进行基因组混组,统计后代中非亲本类型占整个群体的比例,以衡量基因组混组的效果。分别经过5轮循环原生质体融合、循环转化或者循环转导,结果显示,重组程度较高者在后代群体中的比例较低,带有4种标记的后代未出现,带有3种标记的后代最高分别为4.53×10?4、1.64×10?4、4.47×10?3,明显低于文献报道的天蓝色链霉菌中同样实验的结果:带4种和3种标记的后代分别占2.5%、17%。对比上述实验的结果和文献报道的天蓝色链霉菌、乳杆菌基因组混组的结果,并结合计算机模拟循环融合过程,分析后认为:要达到较充分的基因组混组,需要有能够实现微生物细胞间高频重组的操作技术作为基础,重组频率应该不低于10?3～10?2数量级。     

TISSUE CULTURE IN THE PRODUCTION OF NOVEL DISEASE-RESISTANT CROP PLANTS

1. Plant tissue cultures form the basis of a number of techniques which have been developed to effect genetic changes in plants. Progress is being made in the application of these techniques in breeding new, disease-resistant cultivars. 2. It is possible to induce and select for mutants among populations of cultured plant cells. Novel disease-resistant plants of a small number of species have been regenerated from cells selected in culture for their resistance to toxins produced by pathogens, both with and without prior exposure to mutagens. It is not known whether such procedures are widely applicable, and the nature of the genetic changes involved has not yet been determined. 3. The tissues of plant species which are propagated vegetatively are normally genetic mosaics with regard to many characteristics, including resistance to disease. Thus, some of the plants regenerated from cultured cells of such species are more resistant to pathogens than the parent plants. Novel plants produced in this way are already being used in some breeding programmes. 4. Many attempts have been made to modify the genomes of cultured plant cells by means of exogenous nucleic acids. The evidence for integration and replication of this genetic material is equivocal. The technique, therefore, offers no immediate prospects for the development of novel disease-resistant plants, but may be important in the long term as methods are perfected for using plasmids and other agents as carriers of useful genes. 5. Steady advances are being made in producing somatic hybrids of crop plants by fusion of isolated protoplasts. In the long term it may be possible to use protoplast fusion to transfer desirable disease-resistance traits between related species which cannot be hybridized by conventional breeding methods. 6. The culture of excised embryos may be used to grow interspecific and inter-generic hybrid plants in cases where incompatibility occurs after normal fertilization. The technique is already being used by breeders in the production of disease-resistant hybrids of crop species. 7. It is concluded that tissue culture has a limited but useful role to play in the development of novel disease-resistant crop plants.     

灵芝麦角硫因高通量检测方法研究

麦角硫因(ergothioneine,EGT)基于其强抗氧化及体内消耗率低的特点,在人体内拥有多种重要的生理功能。但是在高产EGT微生物筛选和新菌株的选育研究中,现有的EGT检测方法因其步骤繁琐、使用的试剂和设备昂贵而亟待改进。本研究基于EGT的理化性质,建立了EGT-硫氰酸铁高通量快速检测体系,同时选用不同EGT产量的灵芝菌株以及灵芝融合新菌株对该检测体系的准确性进行验证。结果表明该方法可以快速准确地比较出样本间EGT产量高低,使原本需3~4天的工作缩短至2~3 h,HPLC验证结果显示,EGT-硫氰酸铁高通量快速检测体系效果良好,体系稳定。本研究结果将为高产EGT微生物的高通量筛选及高产EGT新菌株的选育提供新的方法和思路。