Optimization of fluorescence in situ hybridization (FISH) for the identification of two polar coccoid green algae species

Standard FISH protocols using fluorochrome-labeled oligonucleotide probes have been successfully applied for in situ detection.However,optimized protocols of FISH for specific eukaryotes in marine environments are often not developed.This study optimized the conditions of fluorescence in situ hybridization （FISH） by using two polar isolated microalgae.The modified conditions were as follows：（1） 10 mg·mL^（-1） lysozyme solution pretreatment at 37℃for 30 min;（2） the hybridization buffer including 20%formamide;（3） the hybridization condition was 47℃for 6 h.The cells enumerated by FISH were compared with those enumerated by flow cytometry（FCM） and DAPI to confirm the cell loss and hybridization efficiency.The optimized protocol was also successfully applied to Arctic Ocean samples,which were found to be dominated by Micromonas sp.The modified protocol showed a high relative efficiency and could be successfully applied for the detection of specific microbial eukaryotes in environmental samples.

DETECTING EXPRESSION OF MRP-1/CD9 mRNA IN LUNG CANCERS USING TISSUE MICROARRAYS AND FLUORESCENCE IN SITU HYBRIDIZATION METHODS

Objective： The aim of this study was to investigate the MRP-1/CD9mRNA expression in lung cancer and normal lung tissues and the relationship between its expression and pathologic grades, clinical stages, metastasis and prognosis. Methods： To observe MRP-1/C9mRNA expression, tissue microarray （TMA） containing 54 lung cancers and 10 normal lung tissues was prepared and Fluorescence in situ hybridization was used. Results： The positive rate of MRP-1/CD9 expression was 48.1% in lung cancer, lower than that of normal lung tissues. The statistical difference was significant （P〈0.05）. Its protein expression had no relationship with the patients＇ ages, sex and the macroscopic type of tumor, but had relationships with the histological type, clinical stage, differentiated degree and metastasis. The expression in non-small cell lung cancer （NSCLC） was higher than that in small cell lung cancer （SCLC）; in well-moderately differentiated group was higher than that in poorly differentiated group; Earlier period group （I＋II） was higher than in later period group （Ⅲ＋Ⅳ）; and in group without lymphoid metastasis was higher than in patients with lymphoid metastasis. Conclusion： The progression of the lung cancer maybe related with the descended MRP-1/Cd9 expression, which may be useful in evaluating the prognosis of cancer patients.

Distribution characteristics of ammonia-oxidizing bacteria in the Typha latifolia constructed wetlands using fluorescent in situ hybridization(FISH)

A molecular biology method, fluorescent in situ hybridization（FISH）, in which the pre-treatment was improved in allusion to the media of the constructed wetlands（CW）, e.g. the soil and the grit, was used to investigate the vertical distribution characteristics of ammonia-oxidizing bacteria（AOB） quantity and the relation with oxidation-reduction potential（ORP） in the Typha latifolia constructed wetlands under three different Ioadings in summer from May to September. Results showed that the quantity of the AOB decreased in the Typha latifolia CW with the increase of vertical depth. However, the AOB quantity was 2-4 times the quantity of the control in the root area. Additionally, ORP in the rhizosphere was found to be higher than other areas, which showed that Typha latifolia CW was in an aerobic state in summer when using simulated non-point sewage at the rural area of Taihu Lake in China and small town combined sewage.

Literature Analysis on Fluorescence in situ Hybridization in China during 2002-2016

In order to explore researches about the chromosome karyotype analysis and fluorescence in situ hybridization(FISH) technology in China,using the bibliometric method,taking " fluorescence in situ hybridization(FISH) " and " chromosome" as key words,this paper made a statistical analysis on the literature published in China National Knowledge Infrastructure(CNKI) during 2002-2016.The results indicated that the number of papers published in 2002 was the smallest(37),while the number of papers published in 2012 was the largest(125).In terms of the distribution of organizations of authors,in 1201 papers,11 organizations published papers ≥15,accounting for 21.65%.In terms of distribution of papers published by different periodicals,11 periodicals published papers ≥10,accounting for 17.65%.In terms of the papers supported by foundation projects,in all papers searched,377 papers were supported by foundation projects,accounting for 31.39%.In terms of the distribution of doctoral and master's dissertations,259 papers were master's dissertations,accounting for 21.57%;92 papers were doctoral dissertations,accounting for 7.66%.

The Cloning and Fluorescence In situ HybridizationAnalysis of Cotton Telomere Sequence

Telomeres form the ends of eukaryotic chromosomes and serve as protective caps that keep chromosomes structure independency and completeness. The first plant telomere DNA was isolated from Arabidopsis thaliana and was shown to have tandemly repeated sequence 5-TTTAGGG-3： The Arabidopsis-type telomere has been found in many plants, but several reports indicate that this sequence is absent in some plants. Up to now, no research has been conducted on the telomere of cotton. In this paper, the Arabidopsis-type telomere sequence was amplified and cloned using the primers designed based on the fragment containing telomere sequence in an Arabidopsis bacterial artificial chromosome （BAC）. Fluorescence in situ hybridization （FISH） with cotton metaphase chromosomes using the Arabidopsis-type telomere sequence as probes indicated that the signals were located at all chromosome ends of seven diploid and two tetraploid cotton species with different signal intensities among chromosome complements of different cotton species, even between long and short arms of the same chromosome. To identify the signals of FISH, the genome DNA of Xinhai 7, a cultivar of Gossypium barbadense, digested by BAL-31 nuclease was introduced in this study. The result of BAL-31 digestion indicated that the hybridization signals of FISH represent the outermost DNA sequence of each cotton chromosomes. So we first proved that the telomeric repeats of cotton cross-hybridize with that of Arabidopsis. The results of terminal restriction fragment （TRF） showed significant variation in telomere length among cotton species. The telomere length of cultivated cotton was close to 20 kb and was larger than those of wild cotton species whose telomere length rahged from 6 to 20 kb.

Using Fluorescence in situ Hybridization to Identify DMD/BMD Deletion Carriers

Objective To identify the deletions in Duchenne/Becker muscular dystrophy (DMD/ BMD) by using fluorescence in situ hybridization (FISH)Methods The exon-specific cosmid DNA probes (representing 18 exons) were used to perform one-color FISH on metaphase and interphase preparations. The peripheral blood samples from 9 normal people (4 males and 5 females) and 5 females from independent deletion DMD/BMD families, as well as 2 amniotic fluid specimens and 2 chorionic villus samples (CVS) from normal pregnant females were analyzed. Results 72%-100% of peripheral blood lymphocyte metaphases or interphases, 60% -70% of amniocyte interphases, and 95 - 99% of chorionic villus cell interphases showed expected signals. One suspected female was identified as deletion carriers and two were excluded.Conclusion FISH in combination with other available techniques allows efficient screening of DMD/BMD deletion carriers, which also lay the ground work for prenatal diagnosis for potential fetal carriers.

Detection of Embryo Sex Chromosome by Dual Color Fluorescent In-Situ Hybridization

In order to evaluate the effects of sex chromosomal mosaicism on the accuracy of single-cell gender diagnosis, sex chromosomes of 21 normal fertilized embryos were detected by dual color fluorescent in-situ hybridization (FISH). The results showed that 4 embryos had sex chromosomal mosaicism (19%) and the remaining 17 showed uniformly XX or XY signals in all blastomeres. In conclusion, identification of sex by dual color FISH analysis of a single cell was accurate and efficient , and sex chromosomal mosaicism would not affect preimplantation gender diagnosis.

Chromosomal mapping of 5S and 18S-5.8S-25S rRNA genes in Saccharina japonica(Phaeophyceae)as visualized by dual-color fluorescence in situ hybridization

It has been reported that there was a linkage of 5S rRNA gene to 18S-5.8S-25S rRNA gene in a few of species in Ochrophyta.In regard to the usual two positions of linked 5S rDNA to the 3′end of 25S rDNA,two pairs of primers were designed for amplification to verify this linkage of two genes in a kelp cultivar of Saccharina japonica,one of species in Ochrophyta.This result supplemented the previous report that 5S rDNA was unlinked to 25S rDNA in this kelp.In order to simultaneously visualize this unlinkage of two genes,dual-color fluorescence in situ hybridization(FISH)technique was applied to the cytogenetics of S.japonica.Dual-color FISH images showed that two and four hybridization signals were present in the kelp gametophyte and sporophyte,respectively,metaphase nuclei hybridized simultaneously with the labeled probes of 18S rDNA and 5S rDNA.Both haploid and diploid karyotypes in decreasing length of chromosomes showed that 18S-5.8S-25S rDNA was localized at the interstitial region of Chromosome 23,whereas 5S rDNA resided at the sub-telomeric region of Chromosome 27.These karyotypes suggested that the kelp nuclear genome had only one locus of each rRNA gene,and their loci on different chromosomes indicated the physical unlinkage of 5S rDNA to 18S-5.8S-25S rDNA in this kelp.Therefore,dual-color FISH seems to be a powerful technique for the discrimination and pairing of chromosomes featured in both small size and nearly identical shape in S.japonica.

粗穗披碱草1H^(t)S染色体特异荧光原位杂交标记开发

小麦-粗穗披碱草1H^(t)S.1BL罗伯逊易位系高抗小麦条锈病和叶锈病,是小麦遗传改良的优异基因源。我们前期利用中国春ph1b基因突变体对该易位系进行诱导,获得了一批诱导后代材料,为了从中准确鉴定小麦-粗穗披碱草1H^(t)S小片段易位系,需要建立能够覆盖粗穗披碱草1H^(t)S染色体的荧光原位杂交(FISH)标记。本研究利用21个小麦及其近缘种探针、61个基于中国春基因组序列新开发的小麦1BS染色体探针以及40个根据简单三碱基重复序列新开发的探针对小麦-粗穗披碱草1H^(t)S.1BL易位系进行非变性FISH分析。结果显示,B74、B76和B77等36个探针(33个为新开发的)在1H^(t)S染色体上具有杂交信号,并且杂交信号可分为仅在染色体末端、仅在着丝粒处、同时在着丝粒处和近着丝粒处、覆盖1H^(t)S约3/4染色体臂、覆盖绝大部分1H^(t)S共五种类型。因此,部分探针单独使用或多个探针联合使用,其信号能覆盖1H^(t)S染色体,能够满足小麦-粗穗披碱草1H^(t)S小片段易位系鉴定,这可为小麦背景中粗穗披碱草染色质追踪提供新的检测手段。

核型分析联合其他遗传学检测技术在高龄孕妇产前诊断中的应用

目的探讨核型分析联合单核苷酸多态性微阵列技术(single nucleotide polymorphism array,SNP array)或荧光原位杂交技术(fluorescence in situ hybridization,FISH)或细菌人工染色体微珠标记技术(BACs on Beads,BoBs)在高龄孕妇产前诊断中的应用价值。方法回顾性研究因高龄行产前诊断的930例孕妇,全部行核型分析,联合BoBs检测420例,联合SNP array检测343例,联合FISH检测167例,分析其染色体结果及妊娠结局。结果930例胎儿中,核型异常192例(20.7%),三体综合征中以21-三体居多,性染色体异常中以克氏征居多。在核型联合其他检测中,联合SNP array检测可显著提高胎儿染色体异常检出率,差异有统计学意义(χ^(2)=6.191,P=0.013)。对于染色体嵌合体的诊断,核型联合FISH检测可更精准地确定嵌合类型及比例。孤立性高龄胎儿染色体异常风险相对较低,高龄合并无创高危胎儿染色体异常率最高,在高龄合并超声软指标中,随超声软指标数目增多,染色体异常率随之升高(P=0.001)。高龄孕妇年龄与核型异常、非整倍体尤其21三体的发生率呈正相关(P<0.05)。结论孤立性高龄孕妇胎儿染色体异常风险相对较低,高龄合并无创高危胎儿染色体异常的风险高,随超声软指标数目增多、孕妇年龄增长胎儿染色体异常的风险增加,核型联合FISH检测可更精准地确定嵌合类型及比例,核型分析联合其他检测尤其与SNP array技术可显著提高染色体异常检出率,有利于遗传咨询及再生育指导,是高龄孕妇首选的产前诊断方案。

Variation of wheatgrass chromosomes in wheat-wheatgrass alien addition line"TAI-27"revealed by fluorescence in situ hybridization(FISH)

The karyotype of the primary wheat wheatgrass alien addition line TAI 27 was 2 n=44 in which all of the chromosomes were metacentric and submetacentric.However,in the progeny of TAI 27 a pair of chromosomes had become small chromosomes in the two morphologically different plants.Fluorescence in situ hybridization(FISH)technique was used to analyze the two different plants.The observations indicate that a pair of small chromosomes in one variation line are from wheatgrass.In another variation line,a pair of small chromosomes are also from wheatgrass,while another pair of wheatgrass chromosomes have substituted the wheat chromosomes.TAI 27 and its variant lines showed a high level of resistance to barley yellow dwarf virus(BYDV).The possible explanation for such a variation and the potential use of the variant lines were discussed briefly.

胡桃楸高质量中期染色体制片及rDNA的物理定位

【目的】以胡桃楸雄花序为材料,建立染色体形态良好、染色体分散且无细胞质背景的胡桃楸高质量染色体制片技术,为深入开展胡桃楸分子细胞遗传学研究提供参考。【方法】通过卡宝品红压片观察花药发育进程,采集花药细胞处于分裂旺盛阶段的雄花序,分别采用1 MPa一氧化二氮(N2O,笑气)、0.7 mmol·L^(-1)环己酰胺、2 mmol·L^(-1)8-羟基喹啉对花序进行不同时间预处理。剥开花序取出花药,酶解后制成悬液,在55℃的烤片机上涂片。以45S rDNA和5S rDNA为探针,对中期染色体进行原位杂交。【结果】1)当雄花序颜色鲜绿、长度达到约1.5 cm、花药顶端变为红色时,花药细胞分裂旺盛,可以观察到大量小孢子母细胞和中期分裂相。因此,为了获得丰富的有丝分裂中期分裂相,取材最佳时机是雄花序中上部的花药顶端变红时,可以保证雄花序中大部分花药处于旺盛分裂期。2)2 mmol·L^(-1)8-羟基喹啉预处理4、6、8 h的染色体都均凝缩不充分,边缘不清晰,拖尾明显,且大部分着丝粒无法辨认;0.7 mmol·L^(-1)环己酰胺预处理4 h的染色体较长,凝缩不充分,拖尾严重,预处理6 h的染色体凝缩适当,形态清晰,大部分着丝粒可以辨认,预处理8 h的染色体边缘清晰,但凝缩过度,着丝粒不明显;1 MPa N2O预处理2 h的染色体长度适中,但凝缩不充分,边缘模糊,处理3、4 h的染色体凝缩程度都较高,染色体较为粗短,染色体间形态差异不明显;此外,N2O预处理4 h的染色体中,有部分会出现粘连拉丝的情况。因此,胡桃楸雄花序最合适的预处理条件为0.7 mmol·L^(-1)环己酰胺处理6 h。3)胡桃楸染色体数为32,染色体基数为16(2n=2x=32)。45S rDNA和5S rDNA探针在胡桃楸染色体上均产生明亮的FISH信号。45S rDNA的杂交信号位于1对中着丝粒染色体的着丝粒附近,2条染色体上的信号强度相近;5S rDNA的杂交信号位于另外1对中着丝粒染色体的着丝粒附近,2条染色体上的信号强度一强一弱。【结论】以胡桃楸花药顶端变红的雄花序为材料,成功建立胡桃楸高质量中期染色体制备及FISH技术体系,为核桃属植物的分子细胞遗传学研究奠定基础,也为花药量大的植物获取高质量染色体制片提供了参考。

微生物荧光原位杂交(FISH)实验技术

为更好地应用荧光原位杂交(FISH)实验进行微生物群落生态研究工作,对FISH实验原理和影响FISH实验结果准确性的主要因素进行分析和讨论,包括寡核苷酸探针的选择与标记、样品的预处理、杂交液和杂交条件的选择等步骤.总结了一些实验心得,提出关键步骤的操作要点和注意事项,以及FISH技术与其他实验手段相结合的发展前景.建议根据不同检测对象和实验目的,对关键步骤进行改进可以提高实验准确性、简化操作和降低成本.

赖草基因组重复序列组成及染色体分布特性

【目的】赖草属植物是麦类作物遗传改良和育种的重要基因资源,但作为异源多倍体植物,其基因组来源仍存在较大争议。通过比较赖草、大赖草及新麦草基因组中重复序列分布,探索赖草属物种基因组来源以及种间基因组多样性的形成特性。【方法】通过构建赖草属物种赖草的Cot-1 DNA文库获得大量重复序列,利用荧光原位杂交技术和重复序列对赖草以及近缘物种大赖草和祖先供体物种新麦草进行染色体荧光原位杂交涂染。【结果】(1)根据序列及基因组分布特性,赖草Cot-1 DNA可归为串联重复序列、散布重复序列、散布加串联混合重复序列以及未能鉴定类型,4种类型占比分别为32.4%、45.7%、12.4%和9.5%。(2)串联重复序列TaiI-family、Lt1-6、pTa-535和pSc250在不同物种及同一物种不同材料间信号数量存在较大变异,分别为7~20,1~14,17~26,0~24个。(3)10个反转座子序列在所有物种染色体的分布呈现3种方式:在所有染色体上杂交信号集中分布在着丝粒、近着丝粒及间质区;在所有染色体的所有区域都有分布;在大部分染色体上的分布方式与第1种相同,但是部分染色体端部也有分布。2个LTR/Copia序列仅在赖草染色体上有分布,其他序列在不同物种以及不同材料间均有分布,但在信号强度以及部分染色体上的分布方式存在多态性。【结论】赖草属物种中的一些重复序列具有快速进化的特性,支持赖草属物种多倍化过程,并存在散在重复序列向整个核基因组的快速同质化扩散。

应用双色间期FISH技术中宫颈脱落细胞取材及制片方法的探讨

目的在双色间期荧光原位杂交技术(FISH)中探讨采集宫颈脱落细胞样本及制片的最佳方法。方法采集在北京大学人民医院门诊就诊174例患者的宫颈脱落细胞,应用新鲜细胞直接涂片(60例)、生理盐水制片(31例),TCT剩余液体直接制片(37例)、TCT盐水制片(30例)和TCT低渗制片(16例)五种取材制片方法;并应用双色间期FISH法,检测宫颈脱落细胞的TERC基因表达率,对比取材制片的最佳方法。结果在直接涂片法、盐水制片法、TCT制片法、TCT盐水制片法和TCT低渗法五种制片过程中,TERC基因的杂交成功率分别是55.0%、61.3%、37.8%、53.3%和81.3%,其中TCT低渗法杂交成功率与TCT制片组比较,有显著差异(P<0.05),与其他方法比较,无明显差异(P>0.05);5种制片方法的玻片背景清晰比例分别为38.3%、54.8%、35.1%、60%和75%,各组之间有明显差异(P<0.05);5组的无/少信号比例分别为18.3%、16.1%、37.8%、23.3%和18.8%,其中新鲜制片的直接涂片法和盐水制片法与TCT直接制片法比较,有明显差异(P<0.05),与其他方法比较无差异(P>0.05)。结论在应用双色间期FISH技术检测宫颈脱落细胞TERC基因时,以TCT低渗制片和新鲜细胞盐水制片操作方法简单快速,杂交成功率最高。

黄瓜倍性材料创制及染色体组成的FISH鉴定

【目的】黄瓜(Cucumis sativus L.)遗传基础狭窄,种质资源多样性较为有限,遗传育种研究相对落后。本试验旨在创制整倍体和非整倍体黄瓜种质材料,建立其准确的染色体组成鉴定方法,为进一步选育黄瓜各种染色体系、目标性状的染色体定位及遗传育种研究奠定基础。【方法】以华北生态型黄瓜‘长春密刺’的高代自交系为材料,0.4%秋水仙素溶液处理萌动种子,诱导染色体数目加倍。为获得同源三倍体材料,以诱导获得的同源四倍体为母本,二倍体为父本进行杂交,授粉35—45 d后采收成熟果实进行胚拯救。采用染色体计数,结合形态学、叶片气孔电镜观察,对诱导株及杂交后代的倍性进行鉴定。利用染色体特异的探针进行荧光原位杂交(fluorescence in situ hybridization,FISH),通过观察特异探针在染色体上杂交信号的数目、强弱及位置,结合黄瓜的染色体形态参数,对诱导株的染色体组成进行鉴定。【结果】对经秋水仙素处理的‘长春密刺’材料进行有丝分裂中期染色体计数观察,结果显示诱导获得8株四倍体(2n=28),3株非整倍体(2n=16,19,27)材料。将四倍体与二倍体杂交获得了三倍体材料(2n=21)。经荧光原位杂交分析,根据黄瓜着丝粒探针Type III和核糖体45S rDNA两类信号在染色体上的信号特征可以看出,与二倍体相比,三倍体与四倍体上杂交信号为倍性变化关系,进一步验证创制出的整倍性材料为三倍体与四倍体。不同倍性‘长春密刺’植株的形态学特征存在一定差异,四倍体植株的形态指标与二倍体差异显著;三倍体植株与二倍体在形态学上差异不显著;非整倍体植株与二倍体在形态学上差异也不显著,但其长势较二倍体弱,且花期推迟,雌雄花花期不遇,坐果率明显低于二倍体。经叶片气孔电镜观察,‘长春密刺’二倍体、三倍体与四倍体植株叶片气孔的大小与密度均存在差异,随着倍性提高,气孔的长度和宽度增加,而气孔密度则下降,说明形态学筛选和叶片气孔电镜观察可以作为鉴定黄瓜倍性的辅助方法。以上述两类黄瓜重复序列(Type III和45S rDNA)和染色体特异的单拷贝基因Csa006700为探针,对染色体数目为16的一株非整倍体诱导株进行染色体组成鉴定。重复序列的荧光原位杂交结果显示,额外的两条染色体为1号或2号染色体。进一步利用黄瓜2号染色体端部的基因Csa006700探针检测,发现该基因只在其中一对染色体上有信号,由此明确该材料为附加两条1号染色体的四体材料(2n=14+2)。研究表明秋水仙素不仅可直接诱导出同源多倍体,同时可诱导各种非整倍体植株。【结论】利用秋水仙素处理黄瓜萌动种子,诱导染色体倍性的变化,结合染色体特异探针的荧光原位杂交鉴定,可快速创制并筛选出各种染色体组成的特异新种质。

棉花细菌人工染色体的荧光原位杂交(BAC-FISH)技术

细菌人工染色体荧光原位杂交(BAC-FISH)技术是植物染色体识别、物理作图等分子细胞遗传学研究的重要工具,但对于某些物种尤其是多倍体植物,由于大量重复序列的存在等问题,使得该技术应用受到很大的限制.通过选择棉花分子遗传图中高重组区的微卫星位点(simple sequence repeats,SSR)标记的策略,筛选到不含或含有少量重复序列的细菌人工染色体(BAC)克隆,同时,在通用FISH技术程序基础上,通过改进发根、变性、洗脱条件等步骤,构建出适合于棉花的BAC-FISH技术,简化了操作流程的同时,获得稳定的杂交结果及较高的检出率;并通过将一随机获得的BAC进行染色体的物理定位,进一步引入双探针、双色及重复杂交技术,显示了该技术的成熟与良好的应用前景和价值.

FISH技术在微生物生态学中的研究及进展

分子生物学技术在微生物生态学研究中具有灵敏、精确和快速的优势,但不能提供微生物的形态学、数量性状、空间分布等信息。荧光原位杂交技术结合了分子生物学的精确性和显微镜的可视性信息,可以在自然生境中监测和鉴定不同的微生物个体,尤其是对难培养和未被培养的微生物进行检测。荧光原位杂交技术被广泛用于微生物群落结构诊断和评价,现已成为微生物分子生态学研究中的热点技术。对荧光原位杂交技术的发展和在微生物分子生态学中的应用进行了综述,探讨了该技术应用中存在的问题和发展前景。

25S rDNA和5S rDNA在大白菜中期染色体上的FISH定位

【目的】确定rDNA在中国大白菜基因组中的位点数目和分布位置,并建立识别大白菜不同染色体的特异标记。【方法】用荧光原位杂交技术对25S rDNA和5S rDNA在大白菜有丝分裂中期染色体进行了定位研究。【结果】在大白菜中期染色体上,分别检出了5对25S rDNA杂交信号和3对5S rDNA杂交信号。对应于大白菜中期染色体形态图,确定5对25S rDNA信号分别分布在大白菜1号染色体长臂(1L)的近末端,2号染色体长臂(2L)的近中部,3号和4号染色体长臂(3L、4L)的近着丝点处和10号染色体的随体上,信号强度为10号>2号>3号和4号>1号;而5S rDNA的3对信号分别位于2号染色体的长臂(2L)和9号、10号染色体的短臂(9S、10S)。【结论】在分子水平上为大白菜部分染色体提供了识别标记。

应用FISH法对污水生物处理系统中细菌数量的计数

近年来,FISH(Fluorescence in Situ Hybridization,荧光原位杂交)法作为一种应用分子生物学技术对细胞及细菌等微生物进行定性以及定量分析的研究方法,在国际上得到广泛的应用。本文介绍了应用FISH法对污水处理工艺中细菌数量的计数方法。同时介绍了活性污泥试样制作、FISH法观测试样的制作以及各种试剂的配制、作用以及使用方法,基因探针的选择原则、杂交方法等实际操作方法与过程。