New cell separation technique for the isolation and analysis of cells from biological mixtures in forensic caseworks

Aim To isolate mucosal cells of the perpetrator in a sexual assault case from a complex mixture of his mucosal cells and the victim’s skin by micromanipulation prior to genomic analysis. Methods To capture and analyze mucosal cells we used the micromanipulation with on-chip low volume polymerase chain reaction (LV-PCR). Consensus DNA profiles were generated from 5 replicate experiments. Results and conclusions We validated the use of micromanipulation with on-chip LV-PCR for genomic analysis of complex biological mixtures in a fatal rape case. The perpetrator’s mucosal cells were captured from nipple swabs of the victim, and a single-source DNA profile was generated from cell mixtures. These data suggest that micromanipulation with on-chip LV-PCR is an effective forensic tool for the analysis of specific cells from complex samples

Complete mitochondrial genomes of Taenia multiceps, T. hydatigena and T. pisiformis: additional molecular markers for a tapeworm genus of human and animal health significance

<p>Abstract</p> <p>Background</p> <p>Mitochondrial genomes provide a rich source of molecular variation of proven and widespread utility in molecular ecology, population genetics and evolutionary biology. The tapeworm genus <it>Taenia </it>includes a diversity of tapeworm parasites of significant human and veterinary importance. Here we add complete sequences of the mt genomes of <it>T. multiceps</it>, <it>T. hydatigena </it>and <it>T. pisiformis</it>, to a data set of 4 published mtDNAs in the same genus. Seven complete mt genomes of <it>Taenia </it>species are used to compare and contrast variation within and between genomes in the genus, to estimate a phylogeny for the genus, and to develop novel molecular markers as part of an extended mitochondrial toolkit.</p> <p>Results</p> <p>The complete circular mtDNAs of <it>T. multiceps</it>, <it>T. hydatigena </it>and <it>T. pisiformis </it>were 13,693, 13,492 and 13,387 bp in size respectively, comprising the usual complement of flatworm genes. Start and stop codons of protein coding genes included those found commonly amongst other platyhelminth mt genomes, but the much rarer initiation codon GTT was inferred for the gene <it>atp</it>6 in <it>T. pisiformis</it>. Phylogenetic analysis of mtDNAs offered novel estimates of the interrelationships of <it>Taenia</it>. Sliding window analyses showed <it>nad</it>6, <it>nad</it>5, <it>atp</it>6, <it>nad</it>3 and <it>nad</it>2 are amongst the most variable of genes per unit length, with the highest peaks in nucleotide diversity found in <it>nad</it>5. New primer pairs capable of amplifying fragments of variable DNA in <it>nad</it>1, <it>rrn</it>S and <it>nad</it>5 genes were designed <it>in silico </it>and tested as possible alternatives to existing mitochondrial markers for <it>Taenia</it>.</p> <p>Conclusions</p> <p>With the availability of complete mtDNAs of 7 <it>Taenia </it>species, we have shown that analysis of amino acids provides a robust estimate of phylogeny for the genus that differs markedly from morphological estimates or those using partial genes; with implications for understanding the evolutionary radiation of important <it>Taenia</it>. Full alignment of the nucleotides of <it>Taenia </it>mtDNAs and sliding window analysis suggests numerous alternative gene regions are likely to capture greater nucleotide variation than those currently pursued as molecular markers. New PCR primers developed from a comparative mitogenomic analysis of <it>Taenia </it>species, extend the use of mitochondrial markers for molecular ecology, population genetics and diagnostics.</p

Functional Genetic Diversity and Culturability of Petroleum-Degrading Bacteria Isolated From Oil-Contaminated Soils

In this study, we compared the culturability of aerobic bacteria isolated from long-term oil-contaminated soils via enrichment and direct-plating methods; bacteria were cultured at 30°C or ambient temperatures. Two soil samples were collected from two sites in the Shengli oilfield located in Dongying, China. One sample (S0) was close to the outlet of an oil-production water treatment plant, and the other sample (S1) was located 500 m downstream of the outlet. In total, 595 bacterial isolates belonging to 56 genera were isolated, distributed in Actinobacteria, Firmicutes, Bacterioidetes, and Proteobacteria. It was interesting that Actinobacteria and Firmicutes were not detected from the 16S rRNA gene clone library. The results suggested the activation of rare species during culture. Using the enrichment method, 239 isolates (31 genera) and 96 (22 genera) isolates were obtained at ambient temperatures and 30°C, respectively, from S0 soil. Using the direct-plating method, 97 isolates (15 genera) and 163 isolates (20 genera) were obtained at ambient temperatures and 30°C, respectively, from two soils. Of the 595 isolates, 244 isolates (41.7% of total isolates) could degrade n-hexadecane. A greater number of alkane-degraders was isolated at ambient temperatures using the enrichment method, suggesting that this method could significantly improve bacterial culturability. Interestingly, the proportion of alkane degrading isolates was lower in the isolates obtained using enrichment method than that obtained using direct-plating methods. Considering the greater species diversity of isolates obtained via the enrichment method, this technique could be used to increase the diversity of the microbial consortia. Furthermore, phenol hydroxylase genes (pheN), medium-chain alkane monooxygenases genes (alkB and CYP153A), and long-chain alkane monooxygenase gene (almA) were detected in 60 isolates (11 genotypes), 91 isolates (27 genotypes) and 93 isolates (24 genotypes), and 34 isolates (14 genotypes), respectively. This study could provide new insights into microbial resources from oil fields or other environments, and this information will be beneficial for bioremediation of petroleum contamination and for other industrial applications

Crystal structure of human dynein light chain Dnlc2A: Structural insights into the interaction with IC74

Abstract The human light chain of the motor protein dynein, Dnlc2A, is also a novel TGF-b-signaling component, which is altered with high frequency in epithelial ovarian cancer. It is an important mediator of dynein and the development of cancer, owing to its ability to bind to the dynein intermediate light chain (DIC) IC74 and to regulate TGF-b-dependent transcriptional events. Here we report the 2.1-Å crystal structure of Dnlc2A using single anomalous diffraction. The proteins form a homodimer in solution and interact mainly through the helix a 2 , strand b 3 , and the loop following this strand in each protein to generate a 10-stranded b-sheet core. The surface of the b-sheet core is mainly positively charged and predicted (by software PPI-Pred) to be the site that interacts with other partners. At the same time, the residues 79-82, 88, and 90 of each molecule formed two holes in the core. Residue 89 of each molecule, which is crucial for the DIC binding function of Dnlc2A, is within the holes. On the basis of these observations, we propose that the homodimer is the structural and functional unit maintained by hydrogen bonding interactions and hydrophobic packing, and that the patch of the surface of the b-sheet core is the main area of interaction with other partners. Furthermore, the two holes would be the key sites to interact with IC74

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Image_2_Functional Genetic Diversity and Culturability of Petroleum-Degrading Bacteria Isolated From Oil-Contaminated Soils.PDF

<p>In this study, we compared the culturability of aerobic bacteria isolated from long-term oil-contaminated soils via enrichment and direct-plating methods; bacteria were cultured at 30°C or ambient temperatures. Two soil samples were collected from two sites in the Shengli oilfield located in Dongying, China. One sample (S0) was close to the outlet of an oil-production water treatment plant, and the other sample (S1) was located 500 m downstream of the outlet. In total, 595 bacterial isolates belonging to 56 genera were isolated, distributed in Actinobacteria, Firmicutes, Bacterioidetes, and Proteobacteria. It was interesting that Actinobacteria and Firmicutes were not detected from the 16S rRNA gene clone library. The results suggested the activation of rare species during culture. Using the enrichment method, 239 isolates (31 genera) and 96 (22 genera) isolates were obtained at ambient temperatures and 30°C, respectively, from S0 soil. Using the direct-plating method, 97 isolates (15 genera) and 163 isolates (20 genera) were obtained at ambient temperatures and 30°C, respectively, from two soils. Of the 595 isolates, 244 isolates (41.7% of total isolates) could degrade n-hexadecane. A greater number of alkane-degraders was isolated at ambient temperatures using the enrichment method, suggesting that this method could significantly improve bacterial culturability. Interestingly, the proportion of alkane degrading isolates was lower in the isolates obtained using enrichment method than that obtained using direct-plating methods. Considering the greater species diversity of isolates obtained via the enrichment method, this technique could be used to increase the diversity of the microbial consortia. Furthermore, phenol hydroxylase genes (pheN), medium-chain alkane monooxygenases genes (alkB and CYP153A), and long-chain alkane monooxygenase gene (almA) were detected in 60 isolates (11 genotypes), 91 isolates (27 genotypes) and 93 isolates (24 genotypes), and 34 isolates (14 genotypes), respectively. This study could provide new insights into microbial resources from oil fields or other environments, and this information will be beneficial for bioremediation of petroleum contamination and for other industrial applications.</p