New approaches to facilitate genome analysis

In this era of concerted genome sequencing efforts, biological sequence information is abundant. With many prokaryotic and simple eukaryotic genomes completed, and with the genomes of more complex organisms nearing completion, the bioinformatics community, those charged with the interpretation of these data, are becoming concerned with the efficacy of current analysis tools. One step towards a more complete understanding of biology at the molecular level is the unambiguous functional assignment of every newly sequenced protein. The sheer scale of this problem precludes the conventional process of biochemically determining function for every example. Rather we must rely on demonstrating similarity to previously characterised proteins via computational methods, which can then be used to infer homology and hence structural and functional relationships. Our ability to do this with any measure of reliability unfortunately diminishes as the pools of experimentally determined sequence data become muddied with sequences that are themselves characterised with "in silico" annotation.Part of the problem stems from the complexity of modelling biology in general, and of evolution in particular. For example, once similarity has been identified between sequences, in order to assign a common function it is important to identify whether the inferred homologous relationship has an orthologous or paralogous origin, which currently cannot be done computationally. The modularity of proteins also poses problems for automatic annotation, as similar domains may occur in proteins with very different functions. Once accepted into the sequence databases, incorrect functional assignments become available for mass propagation and the consequences of incorporating those errors in further "in silico" experiments are potentially catastrophic. One solution to this problem is to collate families of proteins with demonstrable homologous relationships, derive a pattern that represents the essence of those relationships, and use this as a signature to trawl for similarity in the sequence databases. This approach not only provides a more sensitive model of evolution, but also allows annotation from all members of the family to contribute to any assignments made. This thesis describes the development of a new search method (FingerPRINTScan) that exploits the familial models in the PRINTS database to provide more powerful diagnosis of evolutionary relationships. FingerPRINTScan is both selective and sensitive, allowing both precise identification of super-family, family and sub-family relationships, and the detection of more distant ones. Illustrations of the diagnostic performance of the method are given with respect to the haemoglobin and transfer RNA synthetase families, and whole genome data.FingerPRINTScan has become widely used in the biological community, e.g. as the primary search interface to PRINTS via a dedicated web site at the university of Manchester, and as one of the search components of InterPro at the European Bioinformatics Institute (EBI). Furthermore, it is currently responsible for facilitating the use of PRINTS in a number of significant annotation roles, such as the automatic annotation of TrEMBL at the EBI, and as part of the computational suite used to annotate the Drosophila melanogaster genome at Celera Genomics

Linear array of conserved sequence motifs to discriminate protein subfamilies: study on pyridine nucleotide-disulfide reductases

BACKGROUND: The pyridine nucleotide disulfide reductase (PNDR) is a large and heterogeneous protein family divided into two classes (I and II), which reflect the divergent evolution of its characteristic disulfide redox active site. However, not all the PNDR members fit into these categories and this suggests the need of further studies to achieve a more comprehensive classification of this complex family. RESULTS: A workflow to improve the clusterization of protein families based on the array of linear conserved motifs is designed. The method is applied to the PNDR large family finding two main groups, which correspond to PNDR classes I and II. However, two other separate protein clusters, previously classified as class I in most databases, are outgrouped: the peroxide reductases (NAOX, NAPE) and the type II NADH dehydrogenases (NDH-2). In this way, two novel PNDR classes III and IV for NAOX/NAPE and NDH-2 respectively are proposed. By knowledge-driven biochemical and functional data analyses done on the new class IV, a linear array of motifs putatively related to Cu(II)-reductase activity is detected in a specific subset of NDH-2. CONCLUSION: The results presented are a novel contribution to the classification of the complex and large PNDR protein family, supporting its reclusterization into four classes. The linear array of motifs detected within the class IV PNDR subfamily could be useful as a signature for a particular subgroup of NDH-2

Detection of humidity-treated aged latent prints using cyanoacrylate fuming and a reflected ultraviolet imaging system (RUVIS)

For the past several decades, challenges in the detection and collection of latent prints exposed to harsh environmental conditions have inspired research in pretreatment methods prior to the application of chemical, physical, or optical-based enhancement techniques. Some of the difficulties associated with processing degraded latent prints are attributed to dehydration, alterations in chemical composition, and physical disturbance of ridge detail. This study seeks to investigate the effectiveness of humidity, cyanoacrylate fuming method (CFM), and a reflected ultraviolet imaging system (RUVIS) on the detection and collection of aged latent palmprints. Prints were exposed to air flow and ultraviolet (UV) light for a period of 0 to 28 days, and subsequently treated with either cool or warm humidity and CFM. RUVIS was then utilized to detect and capture friction ridge detail after each treatment step. Improvements in RUVIS detection between treatments were evaluated based on four response factors: minutiae count, percent print recovery, ridge thickness and contrast. By measuring these factors, each latent print photograph was able to be converted to quantifiable data to facilitate statistical analysis of potential differences or improvements between treatments. The results demonstrate that the application of 80% relative humidity successfully revived aged latent palmprints across all factors. The combined effect of humidity followed v by CFM treatment and RUVIS detection was greatest for minutiae count and ridge thickness, while percent print recovery and contrast demonstrated more modest improvements when compared to control prints. Additionally, cool temperature treatments outperformed warm temperature treatments across all factors except contrast. The data therefore suggest that to achieve print rejuvenation and overall improvements in RUVIS detection, combined cool humidity and CFM is more effective than humidity alone. The data also indicate a potential correlation between temperature treatments and latent print age. Warm humidity combined with CFM appeared to best enhance RUVIS images on fresher prints of a few days to one week old, while cool humidity and CFM appeared to maximally enhance RUVIS images on prints of several weeks old

An Exploration of Protein and DNA Components in Fingerprint Residue

The main focus of this project was to investigate the protein and DNA components in both sebaceous and eccrine fingerprints. This study investigated the relative content of DNA and proteins in eccrine fingerprints to sebaceous fingerprints. All volunteers were instructed to wash and dry their hands prior to depositing parallel thumbprints. Twenty volunteers were instructed to touch their face to produce sebaceous prints, and 5 volunteers were instructed to wear gloves over a heat source to produce sweaty or eccrine prints. Microscopy was used to score the cellular debris of the right fingerprint on a scale of 1-4 based on density of cellular debris. The score was then compared to the DNA yield and proteins detected in the left fingerprint(s). The results of the study illustrated that sebaceous samples had an average DNA yield of 1101.4 ± 1344.0 pg and eccrine samples had an average DNA yield of 131.7 ± 219.5 pg. The difference in DNA yield between the two sample types was significant (p=0.023). The sebaceous samples (n=20) had an average count of 26 ± 22 proteins, and the eccrine samples (n=5) had an average count of 39 ± 20 proteins. The difference in the number of proteins detected was not significant (p=0.153). The sebaceous samples had better STR quality resulting in 75% full profiles compared to the 20% full DNA profiles of eccrine samples. Linear regression results indicate a lack of correlation between cellular debris scores and DNA yields in sebaceous samples (R=0). A strong correlation between the cellular debris and DNA yield in eccrine samples (R=0.80) needs to be confirmed with more samples. Linear regression results show a moderate correlation between the cellular debris and number of proteins detected for sebaceous and eccrine samples (R=0.39, 0.55, respectively). The results of this study provide additional information about donor variability/shedder status, and the content of DNA and proteins in fingerprint samples

Optimizing A Method For Simultaneous Recovery of Proteins and DNA from Fingerprints

DNA testing on touched objects is a valuable tool in forensic investigations, but DNA is usually present in low amounts, causing poor STR typing results. For touch DNA evidence, there is a clear need for additional individualization, especially for highly probative samples. This could be achieved by testing genetically variable proteins. The goal of this project was to develop a DNA/protein co-extraction method to facilitate DNA and protein testing on the same evidence item. Existing DNA extraction methods were carefully adjusted to allow for downstream mass spectrometry analysis. Initial experiments on saliva and fingerprints placed on glass suggested that trypsin and Millipore microcon MW100 units can be used to extract both DNA and protein from forensic samples, as well as produce interpretable DNA profiles and peptide sequences. The Microcon separation of both fractions was more suitable for both STR and mass spectrometry analysis than simply dividing the sample in half after trypsin digestion with no further purification. When tested in parallel to the standard Proteinase K method, the microcon co-extraction method had better DNA typing success rates. Mass spectrometry results for the microcon trypsin-based co-extraction method yielded expected amounts of identified proteins, including tissue specific proteins for both skin and saliva samples

Open Babel: An open chemical toolbox

Background: A frequent problem in computational modeling is the interconversion of chemical structures between different formats. While standard interchange formats exist (for example, Chemical Markup Language) and de facto standards have arisen (for example, SMILES format), the need to interconvert formats is a continuing problem due to the multitude of different application areas for chemistry data, differences in the data stored by different formats (0D versus 3D, for example), and competition between software along with a lack of vendorneutral formats. Results: We discuss, for the first time, Open Babel, an open-source chemical toolbox that speaks the many languages of chemical data. Open Babel version 2.3 interconverts over 110 formats. The need to represent such a wide variety of chemical and molecular data requires a library that implements a wide range of cheminformatics algorithms, from partial charge assignment and aromaticity detection, to bond order perception and canonicalization. We detail the implementation of Open Babel, describe key advances in the 2.3 release, and outline a variety of uses both in terms of software products and scientific research, including applications far beyond simple format interconversion. Conclusions: Open Babel presents a solution to the proliferation of multiple chemical file formats. In addition, it provides a variety of useful utilities from conformer searching and 2D depiction, to filtering, batch conversion, and substructure and similarity searching. For developers, it can be used as a programming library to handle chemical data in areas such as organic chemistry, drug design, materials science, and computational chemistry. It is freely available under an open-source license fro

Recovery of DNA Profiles from Fingerprints on Paper after the Application of Ninhydrin or DFO Given Certain Time Periods

This study examined the recovery of DNA profiles from fingerprints on paper. This examination occurred in three phases: initial determination of DNA profiles on paper, recovery of DNA profiles given certain time periods, and recovery of DNA profiles after the application of the chemical enhancement techniques of ninhydrin or DFO given certain time periods. Phase II and Phase III paper sheets were exposed to the environment to simulate a house setting. All samples from the phases were extracted with phenol-chloroform. Samples from Phase I and Phase II were amplified with AMPF/STRRTM IdentifilerRTM amplification kit while samples from Phase III were amplified with Promega Power Plex RTM 16 amplification kit. Phase I samples contained, on average, the most amount of DNA with 21.05pg/microL. Phase III samples contained, on average, the lowest amount of DNA with .56pg/microL. It was found that no profiles were recovered from the samples after analysis. It is not recommended to perform DNA analysis after the application of ninhydrin and DFO on paper unless all other options have been exhausted