A preliminary evaluation study of new generation multiplex STR kits comprising of the CODIS core loci and the European Standard Set loci.

The GlobalFiler™ (Life Technologies), Investigator® 24plex QS (Qiagen), and PowerPlex® Fusion 6C (Promega) kits are the latest generation 6-dye fluorescent chemistry STR-PCR amplification kits. These kits allow for the simultaneous amplification of the CODIS core loci and the European Standard Set loci, as well as a few Y-STR loci in addition to the standard sex-determining marker Amelogenin. The present study was designed to be a preliminary evaluation of the three STR-PCR kits in terms of sensitivity, profile recovery from degraded DNA samples, tolerance to PCR inhibitors, and detection of minor components in DNA mixtures. The results showed that the three STR-PCR kits had relatively similar performance with each kit faring better for the different aspects studied. The PowerPlex® Fusion 6C and the Investigator® 24plex QS kits were shown to tolerate inhibitors better, while the GlobalFiler™ kit appeared to have a higher mean percentage recovery of alleles from low template DNA samples and for minor components in DNA mixtures.

Mutation of histone H3 serine 86 disrupts GATA factor Ams2 expression and precise chromosome segregation in fission yeast.

Eukaryotic genomes are packed into discrete units, referred to as nucleosomes, by organizing around scaffolding histone proteins. The interplay between these histones and the DNA can dynamically regulate the function of the chromosomal domain. Here, we interrogated the function of a pair of juxtaposing serine residues (S86 and S87) that reside within the histone fold of histone H3. We show that fission yeast cells expressing a mutant histone H3 disrupted at S86 and S87 (hht2-S86AS87A) exhibited unequal chromosome segregation, disrupted transcriptional silencing of centromeric chromatin, and reduced expression of Ams2, a GATA-factor that regulates localization of the centromere-specific histone H3 variant CENP-A. We found that overexpression of ams2(+) could suppress the chromosome missegregation phenotype that arose in the hht2-S86AS87A mutant. We further demonstrate that centromeric localization of SpCENP-A(cnp1-1) was significantly compromised in hht2-S86AS87A, suggesting synergism between histone H3 and the centromere-targeting domain of SpCENP-A. Taken together, our work presents evidence for an uncharacterized serine residue in fission yeast histone H3 that affects centromeric integrity via regulating the expression of the SpCENP-A-localizing Ams2 protein. [173/200 words].

T3SEdb: data warehousing of virulence effectors secreted by the bacterial Type III Secretion System.

BACKGROUND: Effectors of Type III Secretion System (T3SS) play a pivotal role in establishing and maintaining pathogenicity in the host and therefore the identification of these effectors is important in understanding virulence. However, the effectors display high level of sequence diversity, therefore making the identification a difficult process. There is a need to collate and annotate existing effector sequences in public databases to enable systematic analyses of these sequences for development of models for screening and selection of putative novel effectors from bacterial genomes that can be validated by a smaller number of key experiments. RESULTS: Herein, we present T3SEdb http://effectors.bic.nus.edu.sg/T3SEdb, a specialized database of annotated T3SS effector (T3SE) sequences containing 1089 records from 46 bacterial species compiled from the literature and public protein databases. Procedures have been defined for i) comprehensive annotation of experimental status of effectors, ii) submission and curation review of records by users of the database, and iii) the regular update of T3SEdb existing and new records. Keyword fielded and sequence searches (BLAST, regular expression) are supported for both experimentally verified and hypothetical T3SEs. More than 171 clusters of T3SEs were detected based on sequence identity comparisons (intra-cluster difference up to ~60%). Owing to this high level of sequence diversity of T3SEs, the T3SEdb provides a large number of experimentally known effector sequences with wide species representation for creation of effector predictors. We created a reliable effector prediction tool, integrated into the database, to demonstrate the application of the database for such endeavours. CONCLUSIONS: T3SEdb is the first specialised database reported for T3SS effectors, enriched with manual annotations that facilitated systematic construction of a reliable prediction model for identification of novel effectors. The T3SEdb represents a platform for inclusion of additional annotations of metadata for future developments of sophisticated effector prediction models for screening and selection of putative novel effectors from bacterial genomes/proteomes that can be validated by a small number of key experiments.