以早熟白菜苔为实验材料,从其基因组DNA中分离出C0t-1 DNA并用生物素标记作探针,25S rDNA用地高辛标记作探针,对有丝分裂中期相染色体进行双色荧光原位杂交。每对染色体上均显示出了特定的C0t-1DNA荧光原位杂交带型,5对染色体上显示出了25S rDNA荧光原位杂交带型。双色荧光原位杂交证实了C0t-1 DNA与25S rDNA二者具有一致的染色体位置特征,表明基于rDNA及C0t-1 DNA的荧光原位杂交核型分析技术,优于目前普遍采用的只基于rDNA的荧光原位杂交核型分析方法。结合C0t-1 DNA与25S rDNA的荧光原位杂交带型和传统的染色体的形态学标记分析方法及白菜已公布的基于rDNA分布的核型分析结果,创建了一个精确的白菜核型。

WithBrassica campestris cv. Zaoshu baicaitai as experimental material,C0t-1 DNA was isolated from the genomic DNA and labeled with Biotin-Nick as a probe, while 25S rDNA was labeled with DIG-Nick. Fluorescence in situ hybridized to the mitotic metaphase chromosomes ofB. campestris showed the speciifc lfuorescence bands ofC0t-1 rDNA on each chromosome pair. However, the speciifc lfuorescence bands of 25S rDNA were detected on 5 chromosome pairs. The dual lfuorescencein situ hybridization conifrmed theC0t-1 DNA and rDNA had identical chromosome sites. And also the karyotyping technique based on a combination of rDNA and C0t-1 DNA chromosome landmarks was superior to alone one. Based on a combination of rDNA sites,C0t-1DNA lfuorescent bands, chromosome lengths and arm ratios, a more exact karyotype idiogram ofB. campestris was described.

本研究参照GeneBank中禾本科植物大麦5S rDNA序列,利用PCR技术扩增获得新疆7个小麦种5S rDNA部分序列,进一步与禾本科植物大麦5S rRNA序列比对,得到了5S rDNA结构和NTS边界范围。序列分析发现,不同类型小麦5S rDNA序列保守程度不同,其中大片段保守性较高,小片段相对较低,7个小麦种5S rDNA序列均存在不同程度的插入和缺失序列,同种不同类型和不同种5S rDNA非转录间隔区（NTS）长度和存在位置均呈现不同程度差异。利用MEGA4.0软件,采用邻接法（NJ）和最大简约法（MP）构建分子进化树并计算种间遗传距离。以7个小麦种进化关系的分析结果为依据,利用5S rDNA两种类型片段建立了两种不同的亲缘关系分类依据,旨在为新疆7个小麦种亲缘关系分析提供一定理论依据,以期为后期种间同源关系分析,建立序列集合,推导系统进化与发育关系,重建发育史和遗传育种奠定一定理论依据。

Referring GeneBank in barley grasses 5S rDNA sequences, partial 5S rDNA sequences from several accessions of 7 wheat species from Xinjiang were obtained by polymerase chain reaction, the 5S rDNA structure and NTS boundaries were obtained by further alignments with barley grasses 5S rRNA sequence. Sequence analysis revealed that the degree of the nontranscribed spacer of the different unit classes was different. The nontranscribed spacer of long repeat classes was less variable than that of short repeat classes. Different degrees of insertion and deletion in 5S rDNA sequences of 7 wheat species from Xinjiang was presented and the different degree of difference was presented in 5S rDNA non-transcribed spacer (NTS) length and position exists in different repeats of one species and different species. Molecular phylogenetic tree was constructed and genetic distance between species was calculated by using the MEGA4.0 software, the neighbor-joining method (NJ) and the maxi-mum parsi

目的 克隆间日疟原虫河南分离株与湖北分离株红内期18S rDNA,并进行同源性分析.方法 采用PCR方法从间日疟患者血样DNA中扩增间日疟原虫18S rDNA,纯化后与pGEM-Teasy质粒连接,转化大肠埃希氏菌JM109;阳性克隆质粒经双酶切鉴定后,进行序列测定,采用BLAST和MEGA4生物软件分析同源性. 结果 间日疟原虫18S rDNA扩增片段大小为998 bp;阳性克隆重组质粒经双酶切鉴定,与预期结果相符;序列测定结果显示,河南、湖北2分离株间日疟原虫18S rDNA序列完全相同,与GenBank中报道的12株间日疟原虫相同序列进行比对,其同源性均大于99％;用邻位连接法(neigh-bor-joining,NJ)和非加权组平均法(UPGMA)2种方法构建系统发生树发现,河南分离株、湖北分离株与间日疟原虫X13926.1株遗传距离小,同属一个分支.结论 克隆了间日疟原虫河南与湖北分离株红内期18S rDNA,该基因序列在不同地理株间遗传稳定.

Objective To clone and homology analyze the sequences of blood stage 18S rRNA-encoding gene fragment of two P.vivax isolates from Henan and Hubei provinces in China.Methods The 18S rDNA fragments were amplified by PCR from the DNA extracted from two P.vivax infection blood samples.After purification,the gene fragments were ligated with plasmid pGEM-Teasy to construct recombinant plasmids,and transformed into E.coli JM109.Positive clones were identified by double enzymes digestion methods.The sequences of inserted 18S rDNA fragments were finally determined and analyzed with BLAST and MEGA4 biological software.Results The amplified 18S rDNA fragments of two isolates were about 998 bp in length,and the 18S rDNA sequence of Henan isolate was same as that of Hubei isolate.As aligned with the corresponding sequences of twelve P.vivax strains deposited in the GenBank database,the indentity of nucleotides was more than 99％ respectively.Based on the 18S rDNA sequence,phylogenetic analysis with

目的 探讨梗阻性黄疸患者胆汁中的微生物群落结构.方法 应用末端标记限制性片段长度多态性(T-RFLP)和克隆文库分析,对昆明医科大学第二附属医院肝胆胰外科三病区2010年10月至2013年10月期间诊断为梗阻性黄疸患者的胆汁进行细菌培养,培养结果为阴性的患者胆汁再进行微生物群落结构进行分析.结果 共117例患者的胆汁纳入研究,胆汁中细菌16S核糖体DNA (rDNA)阳性率为42.7％ (50/117).不同梗阻原因胆汁中细菌16S rDNA阳性率(97.3％比17.5％)差异有统计学意义(P＜0.05),而梗阻位置相同的细菌16S rDNA阳性率(43.3％比38.1％)差异无统计学意义(P＞0.05).结论 采用T-RFLP方法分析16S rDNA克隆片段能够有效评估梗阻性黄疸患者胆汁中的细菌群落存在和多样性.

Objective To determine the microbial community structure in bile from obstructive jaundice.Methods Structure of bile microbial community from obstructive jaundice patents was investigated by terminal restriction fragment length polymorphism (T-RFLP)and clone libraries approaches.16S ribosomal DNA (16S rDNA) was retrieved from the patients with obstructive jaundice and negative bile culture from October 2010 to October 2013 in our ward.Results 117 cases of patients included in the study of bile.According to percentage of 16S rDNA stone from 50 patients with negative bile culture.The positive rate of bacterial 16S rDNA was 42.7％ (50/117).The positive rate of bacterial 16S rDNA have different reasons for biliary obstruction in a significant difference (97.3％ vs.17.5％),but there is no statistical difference between the position of obstruction (43.3％ vs.38.1％).Conclusion The result illuminated that T-RFLP analysis of cloned 16S rDNA fragments is a powerful tool for estimating

为红树林植物引进优良种质资源,用改良CTAB法分别提取生长于广东珠海淇澳岛红树林湿地公园的3种红树基因组DNA,以真核生物rDNA ITS序列的通用引物分别对其ITS区进行PCR扩增后测序,对测序结果进行比对、分析。结果表明,不同来源红树的rDNA ITS序列存在扩增碱基差异,无瓣海桑、尖叶卤蕨和猫尾木的rDNA ITS序列长度分别为731bp、615bp和707bp,将3种红树植物ITS序列测定结果提交到GenBank,获得了登录号分别为KJ161168,KJ161166,KJ161167。

The genome DNA of three mangrove species from Qi’ao island ,Guangdong was extracted by the modified CTAB method,then the rDNA ITS amplified by PCR with eukaryotic Ponsensus primers was sequenced,and finally the sequencing results were compared and analyzed to provide a reference for introduction of excellent mangrove germplasm resources.There is difference in the amplified base of rDNA ITS sequence among different mangrove species.The rDNA ITS sequence length of Sonneratia apetala , A.speciosum and D.caudafelina is 731 bp,615 bp and 707 bp respectively.The accession number of ITS sequence of S.apetala,A.speciosum and D.caudafelina is KJ161168,KJ161166 and KJ161167 in GenBank separately.