Development and characterization of new allohexaploid resistant to web blotch in peanut

Peanut diseases seriously threaten peanut production, creating disease-resistant materials via interspecific hybridization is an effective way to deal with this problem. In this study, the embryo of an interspecific F_1 hybrid was obtained by crossing the Silihong(Slh) cultivar with Arachis duranensis(ZW55), a diploid wild species. Seedlings were generated by embryo rescue and tissue culture. A true interspecific hybrid was then confirmed by cytological methods and molecular markers. After treating seedlings with colchicine during in vitro multiplication, the established interspecific F_1 hybrid produced seeds which were named as Am1210. With oligonucleotide fluorescence in situ hybridization(Oligo FISH), molecular marker evaluations, morphological and web blotch resistance characterization, we found that: 1) Am1210 was an allohexaploid between Slh and ZW55; 2) the traits of spreading lateral branches, single-seeded or double-seeded pods and red seed coats were observed to be dominant compared to the erect type, multiple-seeded pods and brown seed coats; 3) the web blotch resistance of Am1210 was significantly improved than that of Slh, indicating the contribution of the web blotch resistance from the wild parent A. duranensis. In addition, 69 dominant and co-dominant molecular markers were developed which could be both used to verify the hybrid in this study and to identify translocation or introgression lines with A. duranensis chromosome fragments in future studies as well.     

简单重复序列(AAG)_5在百萨偃麦草染色体上的分布

百萨偃麦草是小麦亲缘物种之一,对环境胁迫和生物胁迫具有很强的抗性,是小麦遗传改良的重要资源。为了对导入的外源染色体及片段进行有效鉴定,利用荧光原位杂交方法,研究了合成的简单重复序列(AAG)5在百萨偃麦草染色体上的分布情况。结果表明,(AAG)5在百萨偃麦草上有多个位点,利用(AAG)5作探针可以将百萨偃麦草的各条染色体区分开来。     

橡胶素基因家族4个成员在橡胶树染色体上的定位

笔者以巴西橡胶树‘热研7-33-97’品种为材料,利用原位PCR技术对巴西橡胶树的4个橡胶素基因(Hev1.1,Hev1.2,Hev2.1,Hev2.2)在染色体的位置进行了物理定位分析,并利用荧光原位杂交技术对原位PCR结果进行了验证。结果表明:Hev1.1,Hev1.2,Hev2.1,Hev2.2基因分别位于巴西橡胶树第8号染色体长臂、第7号染色体长臂、第6号染色体短臂和第12号染色体长臂上;信号距着丝粒平均百分距分别为10.88,31.51,63.81和67.92。     

In-situ enrichment and analysis of atrazine-degrading microbial communities using atrazine-containing porous beads

We examined the community composition of microbes that colonized atrazine-containing beads buried in agricultural soils that differed in atrazine treatment history. Bacterial abundance was 5-40-fold greater in atrazine-fortified beads. In beads containing 20 mg atrazine kg⁻¹ buried in soil with a history of atrazine application (conditioned soil), the abundance of Actinobacteria increased approximately 80-fold whereas in control soil, Actinobacteria were enriched only 10-fold and the gamma-Proteobacteria and Planctomycetes increased by 60- and 25-fold, respectively. The gamma-Proteobacteria were enriched by 120- and 230-fold in beads containing 200 mg atrazine kg⁻¹ in conditioned and control soil, respectively. The results demonstrate that BioSep^® beads are a suitable matrix for recruiting a diverse subset of the bacterial community involved in atrazine degradation.     

Direct detection of <Emphasis Type="Italic">Taphrina deformans</Emphasis> on peach trees using molecular methods

The ascomycetous fungus Taphrina deformans is the agent of peach leaf curl, a worldwide disease of peach potentially devastating to both crop yields and tree longevity. Conspicuous leaf curl symptoms result from the invasion of host tissue by the strictly parasitic mycelial phase of the T. deformans dimorphic life-cycle. Successful isolation of the fungus in pure culture is cumbersome and limited to late spring/early summer (time of ascospore discharge from infected leaves) and only rarely has the asymptomatic yeast phase been isolated from buds. Molecular methods, namely those based on the hybridisation of nucleic acids, are advantageous for diagnostic purposes since they do not require isolation of the fungus on culture media. Direct amplification using the polymerase chain reaction (PCR) and fluorescent in situ hybridisation (FISH) were tested for diagnosis of peach leaf curl disease in order to provide a fast and reliable method for disease risk assessment. Specific primers and probes were designed based on available ribosomal DNA sequence data. Positive and specific diagnoses of peach leaf curl were achieved with primer TDITS1, using PCR-detection, and probe TDE634, using FISH, both on infected leaves and in washings of asymptomatic peach buds.     

Karyotype analysis of Oreochromis mossambicus,O. urolepis hornorum and their hybrid based on Cot‐1 DNA bands by fluorescence in situ hybridization

Chromosomal karyotypes of Oreochromis mossambicus and O. urolepis hornorum and their hybrid were analysed by means of Cot‐1 DNA bandings through fluorescence in situ hybridization (FISH). To identify all chromosomes, Cot‐1 DNA – which contains highly and moderately repetitive DNA – was extracted from genomic DNA, labelled as a probe with Dig‐11‐dUTP, and in situ hybridized to spreads of mitotic chromosomes of the three samples. The hybridized signals were detected by means of Cy3‐conjugated antidigoxigenin. The FISH results indicated that the three samples had the same diploid number (2n=44) of chromosomes. Specific fluorescence signal bands were detected on all individual chromosome pairs. On the basis of Cot‐1 DNA FISH banding patterns and chromosome morphology, the karyotypes of the three samples have been constructed; no remarkable differences were detected between the karyotypes of these species using this method. These results – which are similar to those reported previously, with respect to chromosome number, morphology and Cot‐1 DNA FISH patterns – suggest chromosomal stasis during speciation and hybridization of tilapia (Oreochromis, Cichlidae). Such a molecular cytogenetic procedure, if used in conjunction with other genomic research methods, could facilitate the study of genomic structure and be adapted for chromosome studies of other animal species.     

小麦背景中披碱草部分染色体的FISH鉴定

为了建立小麦背景中粗穗披碱草(Elymus trachycaulus)和纤毛披碱草(Elymus ciliaris)染色体的跟踪鉴定方法,本研究利用寡聚核苷酸Oligo-pSc119.2-1和Oligo-pTa-535-1为探针的双色荧光原位杂交(FISH)和以(GAA)₈为探针的单色FISH,分别对4个小麦-粗穗披碱草附加系和5个小麦-纤毛披碱草附加系染色体进行分析。结果发现,经与小麦FISH核型比对后,建立了可用于追踪小麦背景中粗穗披碱草1S^t、5H^t、6H^t和7H^t染色体的标准FISH核型;而纤毛披碱草S^c和Y^c染色体FISH信号较弱。FISH检测发现,小麦-粗穗披碱草1S^t附加系自交自发变成了1S^t(1BS·3BL)代换系,且在其染色体组中检测到了一对T1BL·3BS易位染色体,同时发现5AS端部寡聚核苷酸Oligo-pSc119.2-1序列发生了删除。另外,在小麦-粗穗披碱草5H^t附加系中发现2B染色体短臂末端删除现象,形成了2B-del和2BS-4AS·4AL易位染色体,而另一条2B和4A为正常的完整染色体。这种染色体结构重排事件表明,部分小麦-远缘物种附加系细胞学并不稳定,因此,繁种前后应对材料进行单株细胞学鉴定。上述小麦-粗穗披碱草染色体结构变异体的获得,为研究染色体结构重排与基因转录表达和表型变化的关系提供了研究基础。     

巴西橡胶树MADS-box基因家族6个成员的染色体物理定位

MADS-box基因家族广泛分布于真核生物中,巴西橡胶树的MADS-box基因家族主要参与花形态建成,对生殖生长起到重要的调节作用。目前,MADS-box基因家族的26个相关基因已被克隆分析,但它们在染色体上的具体位置还未确定。本研究以巴西橡胶树‘热研7-33-97’品种为材料,将MADS-box基因家族的6个成员（HbAGL8、HbAG15、HbAGL30、HbTT16、HbAP1和HbSVP1）定位在细胞核染色体上,通过双探针荧光原位杂交技术（FISH）对巴西橡胶树MADS-box基因家族的这6个成员在细胞核染色体上进行物理定位分析。结果表明：MADS-box基因家族的6个基因分别位于不同的染色体上,其中HbAGL15、HbAG8、HbAG30和HbSVP1基因定位在第4、5、7和8号染色体长臂上,其信号位点到着丝粒的平均百分距离是11.85、39.71、48.94和6.70;HbTT16和HbAP1基因定位在第1和13号染色体短臂上,其信号位点到着丝粒的平均百分距离是22.19和18.01。本研究结果揭示了巴西橡胶树MADS-box基因家族的6个成员在细胞核染色体上的实际位置,展现家族基因之间的分布特点和连锁遗传关系,不仅丰富了橡胶树分子细胞遗传学信息,也为橡胶树的分子辅助育种和比较基因组学研究提供了分子细胞遗传学的科学理论依据。     

内麦系列小麦品种(系)的染色体结构变异分析

为明确内麦系列小麦品种(系)的染色体结构特点,利用荧光原位杂交(fluorescence in situ hybridization, FISH)技术和pTa535、pSc119.2探针对21份内麦系列小麦材料的染色体进行分析。结果显示,5份材料含1RS/1BL易位染色体;探针除3A、7A、2B、3B、4B、2D和4D外的其他染色体在21份材料间存在结构差异;共鉴定出49种染色体多态类型,其中数量最多的是A基因组(22),其次是B基因组(14)和D基因组(13);在5B、6B、3D和7D染色体的同源染色体间存在不对称核型;根据A、B和D基因组的FISH核型,把21份材料分为16类。通过FISH信号模式,发现所有材料在染色体水平上都有变异。多数材料的染色体类型一致,推测这些材料的遗传背景一致或者相似。     

环境生物新技术在土壤微生态研究中的应用

鉴于传统的微生物分离培养技术已不能满足环境微生物研究的需要。为此,在介绍变性梯度凝胶电泳(DGGE)以及荧光原位杂交(FISH)等分子生物学新技术的基础上,对这些技术在土壤微生态研究中存在的问题和发展前景做了展望;提出将这些技术整合应用到复杂土壤的环境微生态研究中,以了解微生物的时空分布、种群结构及其与功能之间的关系。