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Biomedical subjects

M Lorente

Publications and source records attributed to M Lorente.

32 records · Page 2Linked to original sources

PCR-based DNA typing of saliva stains recovered from human skin.

Human bites in cases of homicide, sexual assault, and abuse are often distorted due to the elasticity and curvature of the skin. Physical comparison of a bite mark to a suspect's teeth is sometimes difficult. Saliva, which is usually deposited during biting, can be collected and analyzed to identify the perpetrator. Using simulated bite mark situations in two experimental series, three samples of 40 microL of whole saliva were deposited on the skin of 27 cadavers (at 33 sites) and three samples of 100 microL of whole saliva were deposited on the skin of 5 cadavers (at 12 sites). Saliva was collected using the double swab technique at t = 5 min, t = 24 h, and t = 48 h. DNA was extracted using the modified Chelex method and submitted to PCR-based typing at two short tandem repeat loci. Results indicate that the concentration of DNA in saliva recovered from skin varies as a function of time since deposition. There is a significant decrease in concentration in the first 24 h but the concentration remains stable from 24 to 48 h. The success of PCR amplification is independent of the time since deposition or the concentration of DNA in the saliva sample. Contamination from the DNA of the cadaver was not found in any of the cases studied.

Bites, Human↗

Sequential multiplex amplification: utility in forensic casework with minimal amounts of DNA and partially degraded samples.

Since its introduction, PCR has become a widely-used, routine technique in forensic laboratories. A number of PCR protocols that were developed originally are now being replaced by more powerful approaches, particularly those based on multiplex amplification of short tandem repeat (STR) loci. One alternative from of multiplex PCR amplification, called Sequential Multiplex Amplification (SMA), was designed to amplify a single locus and then recover and reuse the remaining genomic DNA as a template for subsequent PCR. The SMA process could be repeated several times. SMA has proven to be useful in typing genomic DNA contained in stored PCR samples and analyzing samples of limited quality and/or quantity for multiple loci. The efficacy of the use of SMA for actual typing of casework samples permitted typing for a second locus 98.11% of the samples considered; 70.75% were typeable for a third locus, and 16.98% for a fourth locus.

Blood Stains↗

Increasing DNA extraction yield from saliva stains with a modified Chelex method.

Recovery, preservation and analysis of body fluid stains is an important aspect of forensic science. PCR-based typing of DNA extracted from recovered stains is often a crucial method to identify a perpetrator or exclude an innocent suspect. This paper reports an improved method of extracting genomic DNA from saliva stains deposited on human skin in simulated bite mark situations. Results of organic (phenol-chloroform) extraction and Chelex extraction were compared to a modified Chelex method developed by the authors. Modifications include pre-extraction preparation with proteinase K and incubations at 56 degrees C and 100 degrees C plus microconcentration of the solution. Quantification results using the classical Chelex extraction method showed that 31.9 +/- 4.22% of the deposited DNA was recovered, but using the modified Chelex extraction method DNA recovery was increased to 47.7 +/- 6.90%. The quantity and quality of extracted DNA was shown to be adequate for PCR-based typing at two STR loci.

Bites and Stings↗

Analysis of short tandem repeat (STR) HUMVWA in the Spanish population.

Amplification by polymerase chain reaction (PCR) of variable number of tandem repeat (VNTR) loci and subsequent typing by electrophoresis and silver staining has become a useful tool for identity testing. One viable group of genetic markers amenable to amplification by PCR is the short tandem repeat (STR) loci. A horizontal discontinuous polyacrylamide gel electrophoresis (PAGE) method was used to type the amplified products of the STR HUMVWA. Typing for VWA of 120 unrelated Spanish Caucasians was done. Six alleles were observed with frequencies in the range 0.096-0.242. The genotype distribution meets Hardy-Weinberg expectations (0.25 < P < 0.50). The heterozygosity was 73.3% and the discrimination power (DP) 0.94. Simultaneously, in a small sample of families (n = 24) no new mutations could be found.

Alleles↗

Sequential multiplex amplification (SMA) of genetic loci: a method for recovering template DNA for subsequent analyses of additional loci.

A method called Sequential Multiplex Amplification (SMA) has been developed whereby a limited amount of DNA extracted from a sample can be reutilized for several single polymerase chain reaction (PCR) amplifications. The method involves recovery of genomic template DNA by microfiltration of PCR-amplified samples. Up to 5 different loci have been typed, each in a single system PCR-based assay, beginning with a test quantity of 5 ng template DNA. Genotypes of the DNA donors were compared with those obtained from individual amplifications and shown to be identical. This could be a useful technique for typing a number of loci from a limited amount of DNa and to recover template DNA from samples previously subjected to PCR. Obviously, when small quantities of template DNA are available, this technique can prove quite useful.

Base Sequence↗

Analysis of the HUMTH01 allele frequencies in the Spanish population.

Genetic marker typing based on DNA amplification by the polymerase chain reaction (PCR) increasingly is being employed in forensic casework and for paternity testing. The allele frequencies were determined using PCR for 120 unrelated Spanish Caucasians for the locus HUMTHOH1. Six alleles were observed, with frequencies ranging from 0.013 (allele 11) to 0.254 (allele 10). The observed heterozygosity was 75.8%, and the power of discrimination is 0.92. The genotype distribution meets Hardy-Weinberg expectations.

Alleles↗

Composite PAGE: an alternate method for increased separation of amplified short tandem repeat alleles.

Genetic typing of short tandem repeat (STR) loci may require electrophoretic separation techniques which, depending on the locus, can resolve alleles differing in size by only 2, 3 or 4 base pairs (bps). Many such loci can be separated by horizontal, discontinuous polyacrylamide gel electrophoresis (hd-PAGE). However, some loci present particular separation challenges. A composite (or step) gel system consisting of 2 zones, anodal and cathodal, which differ by pore size and ionic strength, has been developed to augment resolution of DNA fragments without a need for additional electrophoretic equipment. Depending on pore size, as well as ionic strength, initial migration can be increased or decreased to effect resolution. When compared with hd-PAGE, this composite gel electrophoretic system can result in enhanced resolution of PCR amplified STR alleles.

Actins↗

Postmortem stability of lung surfactant phospholipids.

The postmortem stability of the main phospholipids of lung surfactant-phosphatidyl choline (PC), phosphatidyl ethanolamine (PE), phosphatidyl inositol (PI), phosphatidyl serine (PS) and sphingomyelin (S) in three different deaths; one caused by fresh-water drowning, one by salt-water drowning, and one from a sodium-pentobarbital overdose has been studied. The drug overdose was considered the control because there was no surfactant involvement. The results show the stability of these kinds of lipids in the first 24 h, with a progressive decrease from 48 h on until 96 h, with a significant correlation to the time of P less than 0.01 in most cases.

Animals↗

Polyasplenia, caudal deficiency, and agenesis of the corpus callosum.

Fullana et al. [Am J Med Genet (suppl. 2): 23-29, 1986] reported on 2 sibs with an autosomal recessive syndrome of caudal deficiency and polyasplenia anomalies. We report on a similar patient in which agenesis of the corpus callosum (ACC) was also found. Such an association has not been reported previously. This finding of ACC is to be interpreted as another midline anomaly rather than as a causally independent malformation.

Agenesis of Corpus Callosum↗

[Nutritional and immunologic state of patients undergoing heart valve surgery].

A series of 60 consecutive patients treated by heart valve replacement surgery under extracorporeal circulation were reviewed. Poor nutrition was assessed in 28% of these cases, the cardiac index in these patients being significantly smaller than in operated patients with normal nutritional states (2.07 +/- 0.8 as against 2.46 +/- 0.7, p less than 0.01). Furthermore, 58% of those with poor nutrition presented postoperative complications: mediastinitis, wound infection, ventricular arrhythmias or secondary pulmonary infection. One death occurred in this group. Postoperative complications in patients with normal nutritional states developed in only 6% and there were no deaths. Duration of hospital stay was significantly longer in patients with poor nutrition (p less than 0.05). Apart from immunoglobulin levels, all immunologic parameters underwent postoperative modifications related to nutritional state. Physiopathologic hypotheses are proposed as well as a programme for increasing nutritional intake in patients in a precarious metabolic state which predisposes them to complications.

Adult↗