Characterization of protoplasts prepared from the edible fungus, Stropharia rugoso-annulata

Protoplast preparation and regeneration conditions of the edible fungus, Stropharia rugoso-annulata Farlow apud Murrill were studied, and the regenerated progenies were characterized in this study. The optimal condition for protoplast preparation was incubation of young mycelia with gentle shaking in 1.5%(w/v) Lywallzyme at 30 °C for 3 h. PGPM (potato/glucose/peptone/mannitol) was the most suitable regeneration medium. Served as osmotic stabilizer, sugars (mannitol and sucrose) were better than inorganic salts (MgSO₄) for clone development and growth. Pre-incubation of protoplasts in liquid regeneration medium resulted in a significantly decreased regeneration rate. Both dikaryotic isolates and monokaryotic isolates could be identified from protoplast-regenerated progenies, with a much higher frequency of monokaryotic isolates identified from the early-developed and fast-growing regenerated clones. Two parental mating types were also identified from protoplasted monokaryotic isolates, but not segregated by 1:1. The mycelial growth rate of protoplasted monokaryotic isolates showed a mating type-dependent model when cultured at different incubation temperatures and pH values, with A2B2 mating type monokaryotic isolates growing faster than those of A1B1 mating type monokaryotic isolates.     

RAPD技术在果树遗传育种研究中的应用

介绍了RAPD技术在果树品种鉴定和分类、系谱分析、构建遗传图谱、杂种鉴定与突变体检测等遗传育种研究领域中的应用,并对该技术在使用中存在的问题及应用前景进行探讨。     

白杨派树种亲缘关系的RAPD分析

运用RAPD技术对白杨派中7个树种和3个人工杂交种共计50个无性系的亲缘关系进行分析.结果表明:筛选出的26条随机引物共扩增产生222条谱带,其中200条为多态性谱带,多态性带达90.09%,且种间遗传变异大于种内无性系间的遗传变异.聚类分析结果显示,参试的50个材料可分为2大类:第一类由山杨、响叶杨、河北杨组成;第二类由毛白杨、银毛杨、84K杨、银白杨、银灰杨、新疆杨和银新杨组成;3个人工杂交种与白杨亚派树种的亲缘关系较近;人工杂交种银毛杨与84K杨间亲缘关系最近.     

Comparison of PCR-based molecular marker analyses of Musa breeding populations

Progress in the breeding of plantain and banana has been restricted by the complex genetic structure and behaviour of cultivated polyploid Musa. Genetic improvement has been hindered due to the large amount of space required for growth and maintenance of plant populations, in addition to the long growth cycle and the low levels of fertility and seed viability characteristic of cultivated genotypes. Molecular marker assisted breeding has the potential to dramatically enhance the pace and efficiency of genetic improvement in Musa. This study was conducted to compare different PCR-based marker systems (RAPD, VNTR and AFLP) for the analysis of breeding populations generated from two diverse Musa breeding schemes. All three assays detected a high level of polymorphism between parental genotypes and within progeny populations. As expected, AFLP assays had by far the highest multiplex ratio while VNTR analysis detected the highest levels of polymorphism. AFLP analysis of a full-sib tetraploid hybrid population confirmed previous reports based on VNTR analysis, of a high frequency of recombination during 2n (3x) gamete formation by a triploid plantain landrace. In addition, both VNTR and RAPD analyses of a full-sib triploid hybrid population suggested a high frequency of homoeologous recombination during n (2x) gamete formation by tetraploid hybrids. In general, there was a poor correlation between estimates of genetic similarity based on different types of marker. The implications of these findings for the molecular breeding of Musa crops are discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

中国甘蓝型油菜遗传多样性的RAPD分子标记

本文利用ＲＡＰＤ方法和统计学分析,对我国７省市和国外引进的总计４０份甘蓝型油菜品种的遗传多样性进行了研究。结果表明,４０个品种的甘蓝型油菜存在着广泛的遗传变异,根据ＲＡＰＤ指纹图谱,通过在ＤＮＡ分子水平上的聚类分析可以将它们分为３大类群,反映出这些品种之间的亲缘关系,并对如何引进甘蓝型油菜资源进行了初浅的讨论。     

华南瓜类疫霉的RAPD遗传多样性分析

为了解来自广东和广西的瓜类疫霉的遗传多样性,利用从180条RAPD引物中所筛选出的多态扩增性强、重复性好的12条引物,对分离自两省区的96株瓜类疫霉进行了全基因组DNA遗传多样性分析和指纹图谱构建。通过对供试菌株的RAPD-PCR扩增,共获得135条DNA标记谱带,其中124条为多态性谱带,多态检测率为91.9%。利用NTSYSpc Version2.1软件对供试菌株间的遗传距离进行聚类分析并构建系统树,以遗传相似系数0.81为阈值,将96个供试菌株划分为12个RAPD群,多数分离物之间遗传相似性较低,在DNA水平上存在显著的遗传变异,具有较丰富的遗传多样性。不同地区间菌株的遗传分化程度不同,分离自黄瓜的菌株遗传分化明显高于分离自冬瓜的菌株。RAPD群与菌株地理来源、分离寄主、致病力、交配型及甲霜灵抗性均无明显的相关性。     

Extension of the linkage map in Citrus using random amplified polymorphic DNA (RAPD) markers and RFLP mapping of cold-acclimation-responsive loci

Genetic mapping with RAPD markers has been initiated in Citrus. Reproducible polymorphism of amplified DNA fragments was obtained with approximately half of the 140 random primers tested, revealing 266 segregating loci. These were tested for linkage using 60 BC₁ progeny from an intergeneric cross of Citrus grandis (L.) Osb. x [Citrus grandis (L.) Osb. x Poncirus trifoliata (L.) Raf.]. A core linkage map was constructed that consists of nine linkage groups containing 109 RAPD markers and 51 previously-mapped RFLP and isozyme markers. A further 79 markers that could not be ordered unambiguously because of their allelic constitution were associated with individual linkage groups and are shown in relation to the core map. The core map has a total length of 1192 cM with an average distance of 7.5 cM between loci and is estimated to cover 70–80% of the genome. Loci with distorted segregation patterns clustered on several linkage groups. Individual clusters of loci were skewed in allelic composition toward one or the other parent, usually C. grandis. This relatively-saturated linkage map will eventually be used to identify quantitative trait loci for cold and salt-tolerance in Citrus. As a beginning we have mapped three loci detected by a cold-acclimation-responsive cDNA.     

Determination of genetic relationships among shape Phaseolus vulgaris populations in a conical cross from RAPD marker analyses

Random-amplified polymorphic DNA (RAPD) markers were used to determine genetic relationships among Phaseolus vulgaris breeding populations. Genetic distances were calculated from the distribution of 317 RAPD markers among 8 parents, 10 individuals from 8 cycle-one populations, 10 individuals from 6 cycle-two populations and 10 individuals from 2 cycle-three populations of a conical cross. Genetic distances between populations and parents were consistent with their degree of relationship in the crossing scheme indicating that a RAPD analysis is a sensitive and useful method for categorizing breeding materials according to their genetic similarities. Genetic variation among individuals within populations increased from cycle one to cycle three and variation among populations within the cycles decreased from cycle one to cycle three in the conical cross. The results showed that this crossing scheme can be used to collect the genetic diversity in eight parents into a single plant breeding population. Abbreviations: CBB, common bacterial blight; GD, genetic distance; RAPD, random-amplified polymorphic DNA     

Advances to improve the eating and cooking qualities of rice by marker-assisted breeding

The eating and cooking qualities of rice are heavily emphasized in breeding programs because they determine market values and they are the appealing attributes sought by consumers. Conventional breeding has developed traditional varieties with improved eating and cooking qualities. Recently, intensive genetic studies have pinpointed the genes that control eating and cooking quality traits. Advances in genetic studies have developed molecular techniques, thereby allowing marker-assisted breeding (MAB) for improved eating and cooking qualities in rice. MAB has gained the attention of rice breeders for the advantages it can offer that conventional breeding cannot. There have been successful cases of using MAB to improve the eating and cooking qualities in rice over the years. Nevertheless, MAB should be applied cautiously given the intensive effort needed for genotyping. Perspectives from conventional breeding to marker-assisted breeding will be discussed in this review for the advancement of the eating and cooking qualities of fragrance, amylose content (AC), gel consistency (GC) and gelatinization temperature (GT) in rice. These four parameters are associated with eating and cooking qualities in rice. The genetic basis of these four parameters is also included in this review. MAB is another approach to rice variety improvement and development in addition to being an alternative to genetic engineering. The MAB approach shortens the varietal development time, and is therefore able to deliver improved rice varieties to farmers within a shorter period of time.     

Evaluation of RFLP and RAPD markers in a comparison of Brassica napus breeding lines

RFLP and RAPD markers were evaluated and compared for their ability to determine genetic relationships in a set of three B. napus breeding lines. Using a total of 50 RFLP and 92 RAPD markers, the relatedness between the lines was determined. In total, the RFLP and the RAPD analysis revealed more than 500 and 400 bands, respectively. The relative frequencies of loci with allele differences were estimated from the band data. The RFLP and RAPD marker sets detected very similar relationships among the three lines, consistent with known pedigree data. Bootstrap analyses showed that the use of approximately 30 probes or primers would have been sufficient to achieve these relationships. This indicates that RAPD markers have the same resolving power as RFLP markers when used on exactly the same set of B. napus genotypes. Since RAPD markers are easier and quicker to use, these markers may be preferred in applications where the relationships between closely-related breeding lines are of interest. The use of RAPD markers in fingerprinting applications may, however, not be warranted, and this is discussed in relation to the reliability of RAPD markers.     

A new mechanism in plant engineering: The potential roles of microRNAs in molecular breeding for crop improvement

MicroRNAs (miRNAs) are small, endogenous, noncoding RNAs that negatively modulate the expression of genes by inhibiting translation or by promoting the degradation of target mRNAs. miRNAs are now known to have greatly expanded roles in a variety of plant developmental processes, in signal transduction, and in the response to environmental stress and pathogen invasion. Because of their ability to inactivate either specific genes or entire gene families, artificial miRNAs function as dominant suppressors of gene activity when brought into a plant. Consequently, miRNA-based manipulations have emerged as promising new approaches for the improvement of crops. This includes the development of breeding strategies and the genetic modification of agronomic traits. Herein, we highlight new findings regarding the roles of miRNAs in plant traits, and describe the current miRNA-based plant engineering approaches. Finally, we consider the feasibility of modulating current approaches to address future challenges such as breeding programs to increase crop yield.     

向日葵种质资源的随机扩增多态性ＤＮＡ（ＲＡＰＤ）研究

采用ＲＡＰＤ方法对我国２１个向日葵（Helianthus annuus L.)基因型和１１个国外引进向日葵因型进行分析,构建了它们的方图谱。从８０个随机引物中筛选出的２５个有效引物共产生１８８条ＤＮＡ片段,大小分布在０．２６－１．９８kb之间,其中１６４例带具有遗传多态性,约占总数的８７．２％,平均每个引物扩增的ＤＮＡ带数为７．５２条。３２个向日葵的遗传相似性分析表明,各基因型间的Ｎei氏相似性系数分布在０．３９８７．０８５３１之间,平均相似性系数为０．６２８１。１１个国外向日葵基因型的Ｎei氏相似性系数分布在０．７１３４－０．８５３１之间,平均相似性系数为０．７８２８,说明国外基因型之间的遗传基础比较狭窄。２１个国内向日葵基因型的Ｎei氏相似性系数分布在０．４７５０－０．８２０６,平均相似性系数为０．６４７８,说明国内向日葵基因型之间的遗传多态性较为丰富。通过非加权算术平均数聚类（ＵＰＧＭＡ）的方法,给制了３２个向日葵基因型之间的遗传关系树图。３２个向日葵基因明显地聚成Ａ、Ｂ两大类群。２１个国内向日葵基因型聚成了Ａ１、Ａ２两个亚类,Ａ１组包括：Ｘ１０、陕西向日葵、Ｄ－Ｓ１２、长岭向日葵６、黑２－Ｓ２－２、Ｔ－Ｃ０８、长岭葵花４、白葵３号、ＢＨ－１０、Ｊ－Ｓ－Ｂ１、张掖白子葵、Ｃ１０１－Ｓ４－３Ｓ４等１２个基因型;Ａ２组包括：ＬＫ３０５－Ｓ８、ＬＫ、ＣＳ－７、长岭葵花Ｓ２－Ｓ２、辉南７－Ｓ１－３、辉南、ＣＹ－ＸＸ１９－ＸＸ２、吉葵１１２、吉葵１１６等９个基因型。１１个国外向日葵基因型划分为Ｂ１、Ｂ２两个亚类,Ｂ１组包括ＬＧ１２０２８Ｑ、ＬＧ９０２３Ｒ、ＣＲＮ１４３、ＳＦ９９０３、ＳＦ９９０２、Ｓ－３３２２、ＳＨ３３２、ＳＨ４１、ＳＦＰ９００１、ＣＲＮ１４４５等１０个法国基因型;Ｂ２组织有来自美国的Ｇ１０１一个材料。     

Analysis of the genetic diversity and relationships among and within species of Hippophae (Elaeagnaceae) based on RAPD markers

RAPD markers were used to assess the genetic diversity and inter- and intra-specific relationships of the genus Hippophae L. and to study the correlation between genetic distances and geographic distances among populations of H. rhamnoides ssp. sinensis. The results analyzed by the percentage of polymorphic loci and Shannon information index indicated that a high level of genetic diversity existed both among and within species of the genus Hippophae. In the UPGMA dendrogram, the species or subspecies were clustered into two main groups but not strictly grouped according to sect. Hippophae and sect. Gyantsensis Lian. The multiple regression analysis and Mantel test both indicated a significant correlation between genetic distance and altitude distance among populations of H. rhamnoides ssp. sinensis, and the cluster analysis suggested that the genetic variation among populations of H. rhamnoides ssp. sinensis was linked to their monophyletic origin. Moreover, some degree of genetic differentiation was found among samples collected at different times.     

Improvement of the nutritional quality of legume seed storage proteins by molecular breeding

Legume seeds contain a large amount of proteins and are one of the essential protein sources for humans and animals. However, the protein, in legume seeds is usually poor in sulfur-containing amino acids, and its nutritional value is lower than the protein from animal sources. Recently plant breeding has become available by the introduction of molecular biology, and a technique, called molecular breeding, was applied to the production of legume seeds that contain proteins with high nutritional quality. This review describes the expression of legume seed protein genes and the transformation of legume plants. Approaches to improve the legume seed storage protein will be discussed.     

旱麦草属种质资源的随机扩增多态性DNA（RAPDs）分析

利用３１个１０ｂｐ随机引物对来自我国新疆和中东地区的１１份旱麦草属种质资源材料以及两个普通小麦亲本和１个普通小麦与东方旱麦草的远缘杂种后代进行了ＲＡＰＤ分析。对扩增形成的３２１条谱带进行的研究发现,该属植物在新疆地区具有较丰富的遗传多样性,同时还将１个根据形态性状定名为光穗旱麦草的样品修正为东方旱麦草。研究还揭示出旱麦草属与普通小麦之间有明显的遗传分化,并发现四部体光穗旱麦草的两个基因组分别来自二     

5. The use of genetic resources in breeding and breeding research

The dramatic increase in yields of agricultural crops over the last 40 years in developed countries has been attributed equally to improved genetic components and improved agronomic practices. The success of plant breeding is based partly on an increased understanding of the parameters involved, to a great extent on improved and more efficient methods of selection, to greater use of available genetic diversity and also to advances in a number of related disciplines including plant pathology, biochemistry, agronomy and genetics. Successes and problems associated with using various genetic resources in plant breeding are illustrated with examples from some of the world's major crops, including potatoes, barley and cotton.     

A method to measure genetic distance between allogamous populations of alfalfa (Medicago sativa) using RAPD molecular markers

 Alfalfa (Medicago sativa L.) is a forage legume of world-wide importance whose both allogamous and autotetraploid nature maximizes the genetic diversity within natural and cultivated populations. This genetic diversity makes difficult the discrimination between two related populations. We analyzed this genetic diversity by screening DNA from individual plants of eight cultivated and natural populations of M. sativa and M.  falcata using the RAPD method. A high level of genetic variation was found within and between populations. Using five primers, 64 intense bands were scored as present or absent across all populations. Most of the loci were revealed to be highly polymorphic whereas very few population-specific polymorphisms were identified. From these observations, we adopted a method based on the Roger’s genetic distance between populations using the observed frequency of bands to discriminate populations pairwise. Except for one case, the between-population distances were all significantly different from zero. We have also determined the minimal number of bands and individuals required to test for the significance of between-population distances. Received: 7 July 1997 / Accepted: 28 October 1997     

利用RAPD对稻蝗属昆虫亲缘关系的研究

通过20个随机引物的PCR扩增,得到了日本主要稻蝗的随机扩增多态性DNA（RAPD）图谱,根据扩增结果,计算了种间相似系数和遗传距离,建立了UPGMA系统树。结果表明,分布没有重叠、种间容易交配、能产生杂种的中华稻蝗台湾亚种与小翅稻蝗的亲缘关系最近;分布重叠的日本稻蝗与中华稻蝗台湾亚种、日本稻蝗和小翅稻蝗的亲缘关系较近。小稻蝗与其它3种稻蝗的亲缘关系较远。     

西藏白菜型油菜遗传多样性的RAPD分析

通过利用22个10bp随机引物对来自西藏高原地区107份白菜型油菜种质资源材料的PAPD分析,探讨了西藏白菜型油菜品种之间的遗传分化关系,结果表明：（1）供试的107份材料共产生236条谱带,其中210条谱带有多态性,占88．98％,说明白菜型油菜在西藏高原地区具有较丰富的遗传多样性;（2）根据引物扩增出的DNA指纹图谱,运用UPGMA分析法,在遗传距离为0．078处,可将供试的107份白菜型油菜划分为11个类群,发现来自于同一地区或气候相似区的品种往往聚在一起,表明西藏高原白菜型油菜品种的相似性与其原产地的地理,气候背景密切相关,并在此基础上,结合西藏高原农业发展历史,气候背景以及地形地貌特点和植物地理学,植物区系学,植物进化论等方面的综合分析,提出西藏高原是世界白菜型油菜起源地的观点。