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Biomedical subjects

M S Rodriguez-Calvo

Publications and source records attributed to M S Rodriguez-Calvo.

15 recordsLinked to original sources

A review of the collaborative exercises on DNA typing of the Spanish and Portuguese ISFH Working Group. International Society for Forensic Haemogenetics.

Since 1992 the Spanish and Portuguese Working Group (GEP) of the International Society for Forensic Haemogenetics (ISFH) has been organizing collaborative exercises on DNA profiling with the aim of making progress on standardization and discussing technical and statistical problems in DNA analysis. A total of four exercises (GEP-92 to GEP-95) have been carried out until now. A consequence of these exercises was the creation of a quality control programme in Spain and Portugal in 1995 which was carried out simultaneously with the GEP-95 exercise. The number of participating laboratories increased from 10 in the first exercise (GEP-92) to 19 in the last exercise (GEP-95). Despite this increasing number of participating laboratories, results remained satisfactory. In the last exercises, all the laboratories used PCR-based DNA polymorphisms with an increasing number of markers obtaining good results. SLPs were used by only 30% of laboratories in the last two exercises but the results indicated a good level of expertise in most of these laboratories. The reasons for these successful results are the common use of the EDNAP protocol for SLP analysis and commercially available kits or common sequenced allelic ladders for PCR-based DNA polymorphisms.

Blood Stains

Comparison of different electrophoretic methods for digital typing of the MS32 (D1S8) locus.

Minisatellite variant repeat mapping by polymerase chain reaction (MVR-PCR) is a new and powerful approach for individual identification from human DNA. This method has been successfully applied to the hypervariable human minisatellite MS32 (locus D1S8). In this work, we further investigate this locus using different electrophoretic techniques which include: agarose gels and nonradioactive hybridization, miniaturized polyacrylamide gels, followed by silver staining, and automatic detection of fluorescent labeled alleles. The advantages and disadvantages of each method are described. Also, MVR codes from a Galician population (NW Spain) are provided.

Chromosome Mapping

Population data on the loci LDLR, GYPA, HBGG, D7S8, and GC in three southwest European populations.

Three Southwest European populations: Galicia (NW Spain), a mixed Spanish population from the rest of Spain (outside Galicia), and a population sample from the Coimbra area (Centre of Portugal) have been studied for the Low Density Lipoprotein Receptor (LDLR), Glycophorin A (GYPA), Hemoglobin G Gammaglobin (HBGG), D7S8 and Group Specific Component (GC). The allele and genotype frequencies found have been compared with other previously published data. All loci meet Hardy-Weinberg expectations in the three sampled populations. There was no evidence of association in any of the three population samples, between the five loci studied. No significant differences were found with Caucasian populations, nevertheless, significant differences were observed between our three population studies and the US SW Hispanic and African populations. The AmpliType PM DNA test greatly facilitates DNA testing in forensic laboratories, providing quick results and a good discrimination power from a single test.

DNA

Double- and single-strand conformation polymorphism analysis of point mutations and short tandem repeats.

Single point mutations in small DNA fragments and single unit differences in simple repetitive DNA can be detected as double-strand conformation polymorphisms using polyacrylamide gel electrophoresis, with and without sodium dodecyl sulfate, even at temperatures as low as 3 degrees C. Changes in a single base are distinguished by means of the analysis of the heteroduplexes, and changes in more than two bases can be distinguished in both homoduplexes and heteroduplexes. Polymerase chain reaction (PCR) conditions can be designed not only to amplify homoduplexes, heteroduplexes and single-strand DNA at the same time, but also to focus the analysis on either single-strand conformation polymorphism (SSCP) or double-strand conformation polymorphism (DSCP). DSCP seems to be advantageous in typing DNA polymorphisms or mutations in loci with few variants, but, because it is necessary to have a simple pattern of all possible combinations of the alleles, it is not as advantageous in typing systems with many variants.

Base Sequence

Isoelectric focusing of human hair keratins: correlation with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns and effect of cosmetic treatments.

A new isoelectric focusing (IEF) technique in polyacrylamide gels with 6M urea and 1.5% Nonidet P40 has been developed to characterize human hair samples. The phenotypes demonstrated with this procedure has been correlated with the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns described by other authors. The method described can be applied in the forensic science analysis of a single human hair. Using the same IEF technique we have studied the changes in electrophoretic patterns of cosmetically treated hair. The characteristics of the modifications observed and its utility in forensic science work are also discussed in this paper.

Detergents

Electrophoretic human leukocyte antigen HLA-DQA1 DNA typing after polymerase chain reaction amplification.

An electrophoretic method (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) is described which permits the identification of human leukocyte antigen HLA-DQA1 types and subtypes without using allele-specific oligonucleotide probes and dot-blot methodology. The procedure can be used in miniaturized gels in combination with automated electrophoretic systems. The PhastSystem is particularly recommended since temperature control is essential. HLA-DQA1*01 and DQA1*0301 can be distinguished in homoduplexes and DQA1*01 subtypes, DQA1*0201 and DQA1*0401 in heteroduplexes (in only 5 h including DNA extraction and PCR amplification). Additional variations to those recognized using commercially available dot-blot methods can be provided since this procedure permits the identification of single base-pair substitutions. In addition, this method is faster and less expensive than commercial methods.

Base Sequence

Comparison between isoelectric focusing methods for the detection of orosomucoid phenotypes.

Orosomucoid (ORM) polymorphism was investigated by different methods including isoelectric focusing in acid pH ranges followed by silver staining, print immunofixation of desialyzed ORM, fixation using a lectin from the sea-weed Codium tomentosum, isoelectric focusing followed by immunofixation in miniaturized gels and isoelectric focusing in immobilized pH gradients. Population genetics studies were carried out in Galicia (NW Spain) and two new ORM variants were found.

Alleles

Plasma protein and red cell enzyme groups in Galicia (north west Spain).

Galactose-1-phosphate uridyl transferase (GALT), esterase D (EsD), and plasminogen (PLG) phenotypes were determined by isoelectric focusing in thin-layer polyacrylamide gels (PAGIF) in a random sample from Galicia. Haptoglobins (Hp) were determined by conventional electrophoresis. The following gene frequencies were observed: for GALT: GALTN: 0.930; GALTD1: 0.044; GALTD2: 0.025; for EsD: EsD1: 0.874; EsD2: 0.104; EsD3: 0.021; for PLG: PLG1: 0.800; PLG2: 0.199; for Hp: Hp1: 0.426; Hp2: 0.573. Population data results of all electrophoretic markers typed until now in Galician population are also included.

Blood Proteins