白囊耙齿菌(Irpex lacteus)总RNA提取方法的建立

以白囊耙齿菌等为材料,建立了一种从白腐真菌中提取总RNA的方法.该方法1 h便可从菌丝中提取总RNA;获得的总RNA经紫外分光光度计检测,A260/A280为1.9~2.0,凝胶电泳分析28S rRNA与18SrRNA亮度比约为2;其纯度和完整性能够达到进行分子生物学其他领域研究的要求.     

Synthesis and Photophysical Characterisation of a Fluorescent Nucleoside Analogue that Signals the Presence of an Abasic Site in RNA

The synthesis and site‐specific incorporation of an environment‐sensitive fluorescent nucleoside analogue ( 2 ), based on a 5‐(benzofuran‐2‐yl)pyrimidine core, into DNA oligonucleotides (ONs), and its photophysical properties within these ONs are described. Interestingly and unlike 2‐aminopurine (a widely used nucleoside analogue probe), when incorporated into an ON and hybridised with a complementary ON, the emissive nucleoside 2 displays significantly higher emission intensity than the free nucleoside. Furthermore, photophysical characterisation shows that the fluorescence properties of the nucleoside analogue within ONs are significantly influenced by flanking bases, especially by guanosine. By utilising the responsiveness of the nucleoside to changes in base environment, a DNA ON reporter labelled with the emissive nucleoside 2 was constructed; this signalled the presence of an abasic site in a model depurinated sarcin/ricin RNA motif of a eukaryotic 28S rRNA.     

Serine Palmitoyltransferase (scs1/lcb2) Mutants have Elevated Copy Number of the L-A dsRNA Virus

The microsomal fraction isolated from serine palmitoyltransferase (lcb2/scs1) mutants is enriched in a 90 kDa protein. The protein was identified as the major coat (Gag) protein of the L-A dsRNA virus particles by partial sequencing and by its interaction with anti-Gag antibodies. The total amount of Gag in whole-cell lysates of scs1/lcb2 mutant cells is greater than in wild-type lystes indicating that the enrichment of the protein in the microsomal fraction of scs1/lcb2 mutant cells may result from increased copy number of the L-A dsRNA virus. This is supported by the finding that the mutants also have increased levels of L-A dsRNA. Altered sphingolipid synthesis in the scs1 mutant cells appears to increase the copy number of the L-A viral particles. © 1997 John Wiley & Sons, Ltd.     

Relationship between F-specific RNA phage genogroups, faecal pollution indicators and human adenoviruses in river water

Recent studies have shown the increasing interest of F-specific RNA phage genotyping to identify major sources of faecal contamination in waters. This study, conducted in a river located in an urbanized watershed with recognized anthropogenic influences, was aimed at evaluating the relevance of direct phage genotyping by real-time RT-PCR. One hundred percent of positive results were obtained with a 5 mL aliquot of river water (n = 31). Phage distribution was modified after cultivation, since the ratio of the two most abundant genogroups (II and I) reached 1.51 log₁₀ by direct RT-PCR-based method versus 0.30 log₁₀ after cultivation (n = 8). For the first time, positive correlations between the concentrations of genogroup II, bacterial indicators and human adenoviruses were observed, which may indicate a human faecal pollution. No correlation between genogroups II and I has been revealed. The concentration of genogroup I was only correlated with water turbidity, suggesting an animal pollution coming from upstream after rainfall events. Among the microbiological parameters studied, only genogroup II/genogroup I ratio shows variations occurring in the major sources of faecal pollution.     

Differential persistence of F-specific RNA phage subgroups hinders their use as single tracers for faecal source tracking in surface water

The four subgroups of F-specific RNA bacteriophages (I-IV) have been proposed as potential tracers for faecal source tracking. Groups II and III predominate in human sources while groups I and IV are most abundant in animal sources. The four subgroups of naturally occurring F-specific RNA bacteriophages were identified in different samples by plaque hybridization with genotype-specific probes and the persistence of each subgroup was evaluated. The proportions of the F-specific RNA bacteriophage subgroups were measured in wastewaters, after inactivation in surface waters or after wastewater treatment and in mixtures of wastewater of human and animal origin. Our results indicate that phage groups differ in their persistence in the environment and to different disinfecting treatments. The greater survival of subgroups I and II in treated samples hinders the interpretation of results obtained with F-specific RNA bacteriophages. The phages of subgroups III and IV were the least resistant to all treatments. These results should be considered when using genotypes of F-specific RNA as sole tracers for faecal source tracking.