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Biomedical subjects

T M Clayton

Publications and source records attributed to T M Clayton.

9 recordsLinked to original sources
PubMed1998

Analysis and interpretation of mixed forensic stains using DNA STR profiling.

The use of multiplex PCR and fluorescent dye technology in the automated detection and analysis of short tandem repeat loci provides not only qualitative information about the profile--i.e. which alleles are present--but can also provide quantitative information on the relative intensities of the bands, and is therefore a measure of the amount of amplified DNA. The availability of this quantitative information allows for the interpretation of mixtures in a detailed way which has not been previously possible with many other human identification systems. In this paper we present a simple approach to the resolution and analysis of mixed STR profiles resulting from the testing of mixed biological stains in forensic casework and highlight factors which can affect it. This approach requires a detailed knowledge--gained through a mixture of experiments and validation studies--of the behaviour of each locus within the multiplex systems described. We summarise the available data from previously published experimental work and validation studies to examine the general principles underlying this approach.

Alleles↗
PubMed1995

Identification of bodies from the scene of a mass disaster using DNA amplification of short tandem repeat (STR) loci.

The accompanying paper in this issue describes work conducted during a collaborative effort to identify the victims of a mass disaster that occurred on the 19th of April 1993 near Waco, Texas. The DNA identification programme was also used partly as an exercise to further investigate the robustness and reliability of a recently developed STR quadruplex. The preceding paper provides details of the loci used and also deals with efforts to assess the applicability of STR profiling and its suitability for forensic investigations of this nature. In this paper, we present the results obtained from 61 Waco bodies. Using reference blood samples and family trees 26 positive identifications were made using a 'paternity style' analytical approach. Worked examples, representing a range of casework situations, are used to illustrate the kind of approach taken in interpretation of the data and highlight factors which affected its success. Additionally, we report on the successful application of a PCR-based gender test to 24 of the Waco bodies.

Child↗
PubMed1995

Further validation of a quadruplex STR DNA typing system: a collaborative effort to identify victims of a mass disaster.

The relatively new, PCR-based technique of Short Tandem Repeat (STR) profiling has been used in the identification of the victims of a mass disaster. The analysis relied upon a recently developed multiplex reaction and the use of automated fluorescence technology to simulataneously analyse four tetrameric STR loci. The performance of the 'quadruplex' test was assessed by use of a collaborative study incorporating a blind trial and was demonstrated to be accurate, reliable and robust. Furthermore, the system proved to be highly successful despite the fact that many of the samples from the mass disaster scene were extremely degraded. The high success rate coupled with the discrimination power of the system enabled many severely decomposed human remains to be positively identified.

DNA↗
PubMed1995

Short tandem repeat typing of bodies from a mass disaster: high success rate and characteristic amplification patterns in highly degraded samples.

We have used a PCR-based DNA-typing method, involving the coamplification of four tetrameric short tandem repeat loci, in the analysis of a large number of severely degraded tissue samples taken from the scene of a mass disaster in which bodies were exposed to extreme thermal, physical and chemical insult. Analysis of the amplified DNA in a number of the samples revealed uniquely sized artifact PCR products resulting from the amplification of degraded genomic DNA as well as characteristic patterns in the amounts of PCR products generated from differently sized loci. This system has proved to be very reliable and robust, and we were successful in typing all of the four loci in 66% of the samples tested and at least one locus in 83% of the cases. A PCR-based sex test also proved to be very effective when applied to the degraded samples.

Alleles↗
PubMed1994

The validation of short tandem repeat (STR) loci for use in forensic casework.

A quadruplex reaction has been developed which amplifies the short tandem repeat (STR) loci HUM-VWA31/A, HUMTHO1, HUMF13A1 and HUMFES/FPS. Detection of the PCR products employs denaturing polyacrylamide gels coupled with fluorescent-based technology. This system has been evaluated for use in routine forensic casework and has been shown to be both robust and reproducible. The quadruplex reaction is as sensitive as the commercially available HLA DQ alpha Amplitype typing system and can be used on both degraded and aged material. The problems of environmental contamination have been shown to be limited provided strict procedural practices are followed-i.e. physical separation of sample extraction and amplified products; the use of dedicated equipment such as pipettes; the separation of amplification preparation area. The ability of the system to detect mixtures and the successful analysis of case stains has shown that this system is well suited as a tool for forensic investigation.

Alleles↗
PubMed1992

Global transcription pattern of phi C31 after induction of a Streptomyces coelicolor lysogen at different growth stages.

Using two complementary strategies for low-resolution S1 mapping, the global pattern of phi C31 transcription was studied after induction of thermoinducible phi C31 lysogens of Streptomyces coelicolor A3(2). A complex pattern of early transcripts was seen, with a peak of abundance at about 10 min post-induction. Nearly all of these transcripts were from DNA located to the right of the c (repressor) gene and to the left of the attP site: a region of about 14 kb. Early transcription was also observed immediately to the left of the c gene. The c gene itself was also induced, with an earlier expression peak (about 5 min post-induction). Primary late transcripts were generally relatively long, but degraded. They apparently corresponded to most of the 18 kb region to the left of the c gene. Some shorter and more persistent late transcripts corresponded to DNA close to or overlapping the cos site. Large late transcripts from a region close to the left-hand end of the phi C31 genome showed evidence of processing to more stable, smaller RNA species. A failure of older cultures (more than 12 h old) to be induced productively was correlated with a much longer period of early transcription, reduced late transcription, failure to synthesize a major virion protein, and failure to package phi C31 DNA. Moreover, heat treatment of the older lysogenic cultures did not result in the phi C31-dependent shut-down of host rRNA transcription previously observed for young cultures (Rodríguez et al., Journal of General Microbiology (1986) 132, 1695-1701; Clayton & Bibb, Molecular Microbiology (1990) 4, 2179-2185).

Bacteriophages↗
PubMed1990

Induction of a phi C31 prophage inhibits rRNA transcription in Streptomyces coelicolor A3(2).

A lysogen of Streptomyces coelicolor A3(2) containing a thermoinducible mutant of the temperate phage phi C31 (phi C31 cts1) was used to obtain synchronous phage development. Filter hybridization experiments indicated a marked reduction in rRNA synthesis after prophage induction. S1 nuclease mapping showed that transcription from each of the four promoters of one rRNA gene set (rrnD) was reduced to approximately the same extent, and that inhibition required protein synthesis. Crude preparations of RNA polymerase from induced lysogens had enhanced transcribing activity for phi C31 DNA which was lost upon further purification. The purified preparations were unimpaired in their ability to transcribe from the rrnD promoters in vitro and apparently unchanged in polypeptide composition. The factor(s) responsible for stimulating phage transcription, and possibly for inhibiting rRNA synthesis, may have been separated from the enzyme during purification.

Bacterial Proteins↗