动物性别决定的分子机理及性别鉴定与控制新技术

哺乳动物中位于Ｙ染色体短臂临界区域的ＳＲＹ基因启动雄性性状的发育。该基因在生殖腺脊中的表达可激发下游基因ＭＬＳ的转录,引起缪氏体抑制物和睾酮分泌,促使睾丸组织器官的发育。针对该基因制备特异探针或产该区域片段得到引物对就可用ＦＩＳＨ或ＰＣＲ准确,快速鉴定出植入母体前的胚胎性别,以及对精子筛选分离的结果作出准确评价。与ＰＣＲ相比,ＦＩＳＨ技术更具检测优势。     

Ability of chromosome 4H to compensate for 4D in response to drought stress in a newly developed and identified wheat–barley 4H(4D) disomic substitution line

A spontaneously developed wheat–barley 4H(4D) disomic substitution line was identified cytogenetically using genomic in situ hybridization (GISH), multicolour fluorescent in situ hybridization (FISH) and microsatellite markers. The ability of the barley 4H chromosome to compensate for wheat 4D in response to mild drought stress was also investigated. In the barley cv. 'Betzes' and the 4H(4D) substitution line, mild osmotic stress induced intensive stomatal closure, resulting in reduced water loss through transpiration and unchanged relative water content in the leaves. As the CO₂ assimilation rate remained relatively high, the water use efficiency, which is an important factor associated with drought tolerance, increased extensively under mild osmotic stress in these lines. In the case of the parental wheat genotypes, however, mild drought stress induced less intense stomatal closure and a greater decrease in the CO₂ assimilation rate than in barley or in the substitution line, resulting in unaugmented or reduced water use efficiency. The results demonstrate that genes localised on the 4H chromosome of barley were able to increase the water use efficiency of the wheat substitution line, which is suitable for improving wheat drought tolerance through intergeneric crossing.     

尼罗罗非鱼六个性别相关标记的FISH分析

从尼罗罗非鱼（Oreochromis niloticus）BAC基因文库5个克隆中提取和纯化含有6个性别连锁或相关标记（CLC5，GM204，GM271，GM354，UNH995和UNH104）的重组质粒DNA作为模板，以简并核苷酸为引物，通过PCR制备原位杂交探针。探针用荧光素进行标记，并与尼罗罗非鱼中期相染色体进行荧光原位杂交以确定这些标记在尼罗罗非鱼染色体上的位置和分布。结果显示，这些性别连锁或相关标记都位于尼罗罗非鱼第一对染色体长臂近末端，从分子细胞学角度验证了第一对染色体是尼罗罗非鱼的性染色体。另外由于这些标记的荧光信号在XY个体的2条性染色体上都有，一方面说明这些标记在罗非鱼上还不是性别特异的；另一方面也验证了尼罗罗非鱼的性染色体还处于分化的早期阶段。【中国水产科学，2006，13（4）：525—529】     

Development and application of fluorescence in situ hybridization (FISH) method for simple and rapid identification of the toxic dinoflagellates Alexandrium tamarense and Alexandrium catenella in cultured and natural seawater

ABSTRACT:   The toxic dinoflagellates Alexandrium tamarense (Lebour) Balech and A. catenella (Whedon and Kofoid) Balech produce potent neurotoxins, such as saxitoxin and gonyautoxin and have been mainly responsible for paralytic shellfish poisoning (PSP) in Japan. To prevent a negative effect on the fishery industry, it is necessary to identify these toxic species precisely and rapidly before and during the bloom. In this paper, a rapid and simple protocol of a fluorescence in situ hybridization (FISH) method using ribosomal RNA (rRNA)-targeted probes has been established for identifying the cultured strains and natural cells of A. tamarense and A. catenella . Using the FISH method established in this study, it was possible to identify these toxic species species-specifically and rapidly, within 30 min. The procedure of detection constituted three steps: (i) fixation/dehydration; (ii) hybridization; and (iii) washing; this made the identification simple. Moreover, this method did not require either special techniques or equipment, and the cost for detection was low. The specificity, rapidity, and simplicity of the developed method suggest that it might be useful for routine monitoring of these toxic microalgae.     

内蒙古绒山羊Hoxa7基因在毛囊中的表达

利用原位杂交技术检测Hoxa7基因在毛囊不同生长时期的表达模式,结果表明,Hoxa7基因分别在胚胎期的95 d初级毛囊的内根鞘表达、在115 d次级毛囊的内根鞘和绒干表达、在125 d初级毛囊内根鞘和毛干皮质表达、在兴盛期10月的初级毛囊毛干皮质和次级毛囊的绒干中表达。揭示在毛囊发育中,Hoxa7基因对于毛囊细胞的增殖可能起一定的作用。     

Fluorescent in situ hybridization as an aid to introducing alien genetic variation into wheat

Summary Fluorescent in situ hybridization (FISH) has been used to assess the occurrence and frequency of wheat-alien chromosome pairing in a wheat/Thinopyrum bessarabicum hybrid and in wheat/rye hybrids with different levels of chromosome pairing by examining pollen mother cells at metaphase I of meiosis. The use of FISH to identify the presence and size of alien chromatin in a wheat background is also demonstrated.The value of FISH as an aid to the introgression of alien genetic variation into wheat is discussed.Abbreviations FISH fluorescent in situ hybridization - GISH genomic in situ hybridization - PRINS primer-induced in situ hybridization     

基因组原位杂交技术在植物研究中的应用

基因组原位杂交是以亲本之一的总基因组DNA做探针,另一亲本的基因组DNA做封阻,在荧光原位杂交技术的基础上发展起来的一种染色体/染色质检测技术。在其发展的十几年里,已在植物的基因组研究中发挥了重要的作用。应用这一技术可对多倍体中基因组之间的亲缘关系、基因组组成及起源进行研究;对杂交种中染色体组的组成进行分析;对代换系、附加系和易位系进行有效的鉴定,并对其中的外源染色体或染色体片段的来源、大小、数目及发生位点进行检测和定位。此外,利用基因组原位杂交技术还有助于确定物种间的同源性;研究杂交种中来源不同的染色质在核中的分布;探索B染色体的起源、染色体间的配对、重组、交换等现象。随着基因组荧光原位杂交技术体系的不断发展、完善和改进,其应用范围不断拓展,在植物基因组研究领域中发挥了越来越重要的作用。     

The study of introgressive lines of Triticum aestivum × Aegilops speltoides by in situ and SSR analyses

Fluorescence in situ hybridization (FISH) with a genome‐specific repeat, Spelt1, and wheat simple sequence repeat (SSR) markers were used to analyse the chromosome constitution of two Triticum aestivum×Aegilops speltoides introgressive lines. The lines 170/98i and 178/98i carried one and two subtelomeric regions of Ae. speltoides (per haploid genome), respectively, marked by Spelt1 repeats according to FISH data. SSR analysis detected homoeologous substitution of wheat chromosome 7D with Ae. speltoides chromosome 7S in the lines 178/98i and 170/98i as well as the assumed terminal translocation in the short arm of chromosome 3A in the line 178/98i. Anthocyanin pigmentation of the coleoptiles was found in the lines 170/98i and 178/98i and resulted from the 7S (7D) substitution. It was demonstrated that Spelt1 could be effectively used for the rapid identification (without DNA isolation) of terminal translocations of T. aestivum×Ae. speltoides introgressive lines as well as for further analysis of the stability of the hybrid plants.     

葱的CPD染色和45S rDNA FISH核型分析

为给葱的染色体的识别提供新标记,建立葱的分子细胞遗传学核型,本研究采用去壁火焰干燥法制备了分散且形态良好的葱中期染色体,并进行了CPD（PI和DAPI组合）染色和45S rDNA荧光原位杂交（FISH）,根据葱染色体的形态特征,结合CPD染色和FISH结果,对葱进行了核型分析。CPD染色结果:葱所有染色体臂末端都显示CPD带。FISH结果：有一对45S rDNA位点（在第5对染色体上）。葱的核型公式：2n=2x=16=2sm+12m+2st(SAT)。研究表明：利用CPD染色和45S rDNA FISH,不仅能为染色体识别提供新标记,还能了解染色体GC丰富区的分布,为葱属植物的物种鉴定、系统分类与进化等研究提供DNA分子方面的证据。     

荧光原位杂交(FISH)技术在转基因小鼠检测中的应用

制备转基因小鼠特有的DIG标记探针，应用染色体荧光原位杂交（FISH）技术，对转基因小鼠外周血细胞的遗传性状进行分析，检测其合子类型，以挑选纯合子小鼠用于保种。结果杂合型小鼠61％的细胞有一个荧光信号，而纯合型小鼠约24％的细胞有两个荧光信号，48％的细胞有一个荧光信号，野生型小鼠无荧光信号。因此染色体荧光原位杂交技术可作为检测转基因小鼠遗传特性的一种方法。