Biomedical subjects

A Carracedo

Publications and source records attributed to A Carracedo.

At least 109 records · Page 6Linked to original sources

PubMed1996

Report on the third EDNAP collaborative STR exercise. European DNA Profiling Group.

This report describes an inter-laboratory exercise completed on behalf of the European DNA Profiling (EDNAP) group. The exercise is one in a series designated to identify STR loci which could be used for harmonisation between participating European forensic science laboratories. Participants were asked to identify the alleles present in five bloodstains at the STR loci HUMTHO1 and HUMVWFA31/A. Two of the stains were prepared from mixtures of two different blood samples. There were no special instructions and each laboratory was requested to use the methodology normally employed for crime case investigations. All participating laboratories achieved the same results for both loci. In addition, the laboratories were also requested to report the results obtained from any other loci which would normally be used in crime case investigations. A comparison of these results showed some inter-laboratory variation.

Alleles↗

PubMed1996

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↗

PubMed1996

Effect of environmental factors on PCR-DNA analysis from dental pulp.

This study was designed to observe the results of DNA typing on teeth subjected to aging, different temperatures and various environmental factors. A total of 570 teeth were studied. The study included the analysis of the PCR-based polymorphisms HLA DQA1, D1S80, HUMTH01, HUMFES/FPS and the XY homologous gene amelogenin. In general the best results were obtained with the XY homologous gene amelogenin, followed by the two STRs studied (HUMTH01 and HUMFES/FPS). The small fragment sizes and the method of detection used after PCR amplification are the main factors explaining this fact. In general, teeth submerged in water gave the poorest results. Teeth exposed to outdoor conditions provided better results than teeth buried in sand or soil, but even in these cases good results were obtained. Up to 4 degree C, temperature had only a slight influence on the results. Positive results were obtained in most cases at high temperatures (400 degrees C for 2 min) which are rarely reached in practical casework. Positive typing results for the XY homologous gene amelogenin and the STRs were obtained from teeth 10-30 years old. The usefulness of dental pulp for identification purposes is exemplified in some actual cases.

DNA Fingerprinting↗

PubMed1996

A highly variable STR at the D12S391 locus.

A total of 103 fragments in the STR D12S391 locus were sequenced. 24 different alleles were found which can be grouped into 12 allelic classes based on the total number of repeats. The structure of this compound STR consists of blocks of (AGAT) and (AGAC) repeats with a consensus structure (AGAT)8-17 (AGAC)6-10 (AGAT)0-1. Whereas shorter alleles only have (AGAT) repeats, > 225 bp alleles are more complex, having two motifs (AGAT) and (AGAC). Population data showed that this to be a highly polymorphic STR with a heterozygosity of 0.9. This fact together with its simple structure make this STR very suitable for forensic and genetic purposes.

Alleles↗

PubMed1996

Semiparametric approach to match probability calculations using single locus probes.

A semiparametric approach to match probability calculations using single locus probes has been developed and compared graphically with other standard methods by a one-sample simulation. The density functions obtained using this method are closer to the real distributions than those obtained by conventional approaches. Our method does not need to establish an arbitrary match threshold, which has been a source of problems in practical applications of standard methods. Moreover, it can be adjusted to any particular conditions by setting the experimental error and correlation of each laboratory. To assess the practical performance of this method we carried out a comparison experiment using a sample of 229 individuals analysed in duplicate.

Alleles↗

PubMed1996

Failed PCR amplifications of MBP-STR alleles due to polymorphism in the primer annealing region.

The PCR-based STR system MBP-B (myelin basic protein locus B) has been reported to exhibit a high rate of mutations. Using a newly designed pair of primers we present evidence that this is due to failed amplifications caused by a polymorphism in the annealing region of the reverse primer originally designed. With the new reverse primer described here no exclusions were found (out of 59 mother/child pairs analysed) while one was detected with the old set of primers. The results obtained with both pairs of primers in a random population sample (n = 112) from North Portugal are compared. In this sample 13 individuals typed as homozygotes with the pair of primers originally described, were found to be heterozygous when the amplifications were performed with the new reverse primer. By sequence analysis, a substitution in the reverse primer binding sequence originally described was determined. This substitution is located upstream from the repetition site and consists of G-->A transition. This variation reaches polymorphic frequency and is responsible for the relatively frequent null alleles due to failed amplifications when the previously designed primers are used.

Alleles↗

PubMed1996

Population study of the STRs HUMTH01 (including a new variant) and HUMVWA31A in Catalonia (northeast Spain).

Allele and genotype frequencies for 2 short tandem repeat loci were determined in a population sample from Catalonia (Spain) using the polymerase chain reaction. After denaturing PAG electrophoresis, seven common and one variant (13.3) alleles were identified for HUMTH01 in a sample of 234 unrelated individuals, and seven alleles were found for HUMVWA31A in 162 individuals. No deviation from Hardy-Weinberg equilibrium was found. The observed heterozygosities are 76.92 and 79.62 respectively. The discrimination power determined for the individual loci is 0.928 and 0.937 respectively.

Alleles↗

PubMed1996

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↗

PubMed1995

Allele frequency distribution of four PCR-amplified loci in the Spanish population.

The allele frequency distributions of four VNTR loci amplified by PCR have been studied in a population of 205 individuals from Spain. The loci analysed are D1S80 and three STRs: HUMTH01, HUMFES/FPS and HUMACTBF2 (SE33). The former was visualized in Metaphor agarose gels, and the STRs in sequencing polyacrylamide gels under denaturing conditions which could separate alleles with differences of a single base. This is of particular importance in the HUMTH01 locus, a tetrameric STR in which two alleles (9.3 and 10) were detected differing in a single base. Furthermore, HUMACTBP2 has at least 30 alleles, some of which may vary by as little as one base. At this locus a variation in the allele mobility was observed, depending on the electrophoretic conditions. For this reason, there should be careful consideration before this marker is accepted and validated as a common interlaboratory system. This paper does not include any comparison of the frequencies obtained for this locus with other recent studies. For the rest of the loci, the frequencies found have been compared with other published population studies; they show a degree of difference, particularly in the D1S80 locus. Finally, the systems were tested for Hardy-Weinberg equilibrium, and some statistical parameters of forensic interest were calculated.

Alleles↗

PubMed1995

The use of the STRs HUMTH01, HUMVWA31/A, HUMF13A1, HUMFES/FPS, HUMLPL in forensic application: validation studies and population data for Galicia (NW Spain).

The 5 tetranucleotide short tandem repeats, HUMTH01, HUMVWA31/A, HUMF13A1, HUMFES/FPS and HUMLPL were studied using different electrophoretic methods and PCR amplification conditions in order to optimize the typing conditions. A genetic population study in the population of Galicia was carried out and the allele and genotype frequencies are given. Compliance with the Hardy-Weinberg equilibrium was tested using different statistical parameters, with clear advantages resulting in favor of using the exact test (Guo-Thompson method) instead of conventional chi-square methods. Some statistical parameters of forensic interest (PD, CE, h) were also calculated. There were no mutations found in a total of 73 paternal meioses and 101 maternal meioses. Abnormal electrophoretic mobility was found in the AT-rich STR HUMF13A1 under non-denaturing conditions and, therefore, the use of denaturing conditions is absolutely necessary. No "stutter" bands were found, although double peaks in the HUMFES/FPS system were observed in some samples. The advantage of using automated sequencers with fluorescent technology is also reported.

Base Sequence↗

PubMed1995

Genetic markers: association study in migraine.

Eleven genetic markers were typed in 112 unrelated patients with migraine (50 with aura, 62 without aura) and compared with a random sample of healthy individuals. No significant differences were found for the ABO and Rh systems, acid phosphatase 1, phosphoglucomutase 1, adenosine deaminase, haptoglobin, transferrin, alpha-1-antitrypsin, and D1S80. Strong associations between the group of patients with migraine and group-specific component GC 1F-1F and esterase-D ESD 2-2 phenotypes were observed. These associations raise the possibility that a molecular genetic factor for migraine may exist in or near the Group Component (chromosome 4) and Esterase D (chromosome 13) loci, and represent a first comprehensive step in the eventual localization and isolation of the migraine genes.

Adolescent↗

PubMed1995

Distribution of the HLA-DQA1 polymorphism in the population of the Basque Country (Spain).

Allele and genotype frequencies at the HLA-DQA1 locus were determined in Basque Country populations using PCR methodology and a dot-blot assay with ASO probes. The results showed no deviation from the Hardy-Weinberg equilibrium. No differences were found between the three Basque Provinces, but gene frequencies were significantly different from those of some other Spanish and world populations.

Alleles↗

PubMed1994

Investigation of the STR locus HUMTH01 using PCR and two electrophoresis formats: UK and Galician Caucasian population surveys and usefulness in paternity investigations.

Two hundred unrelated Caucasians from the UK and 210 from Galicia (NW Spain) have been genotyped for the HUMTH01 locus using polymerase chain reaction (PCR) amplification followed by high sieving agarose electrophoresis (UK samples) and polyacrylamide electrophoresis (Galician samples). Allele and genotype frequencies obtained from both populations were in close agreement to those seen by other workers and chi 2 tests of the two populations showed that both were in Hardy-Weinberg equilibrium. The potential usefulness of HUMTH01 in paternity investigations was analysed by constructing, within each population, false family trios, each with a non-father, in order to estimate the actual exclusion rate. The observed rate of exclusion was in close agreement with the expected rate for both populations. Examination of the mother-child pairs in the false families showed a common allele in every case and no evidence of mutation or non-Mendelian inheritance was observed. The HUMTH01 locus shows an informative polymorphism and the production and analysis of population databases is easily achieved. The use of PCR followed by electrophoresis in either gel format appears to provide a quick and straightforward method for investigating the HUMTH01 locus.

Adult↗

PubMed1994

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↗

PubMed1993

The distribution of HLA DQA1 and D1S80 (pMCT118) alleles and genotypes in the populations of Galicia and central Portugal.

Two South-West European populations (Galicia and Central Portugal) have been studied for the HLA DQA1 and D1S80 systems. The allele and genotype frequencies found have been compared with other previously published data. The distribution of the observed genotypes is in Hardy-Weinberg equilibrium for both systems. In the D1S80 system, no significant differences were found between both populations, although in the HLA DQA1 system the allele DQA1*0301 is twice as frequent in the Galician population. Other populations that have been compared showed a certain degree of divergence for the HLA DQA1 system. The combined chance of exclusion for both systems is 0.84 in Galicia and 0.85 in Central Portugal, and the combined power of discrimination is 0.993 in the 2 populations studied.

Alleles↗

PubMed1993

Population genetics of three VNTR polymorphisms in two different Spanish populations.

Two different Spanish populations, one from Galicia (NW Spain) and the other from the rest of Spain, have been analyzed at three different hypervariable loci (YNH24, MS43a and MS31) using the EDNAP electrophoretic protocol and HinfI as restriction enzyme. Although the "rest of Spain" population is a clearly stratified population using classical blood groups, no evidence of stratification for these loci has been found and the differences to the Galician population were not significant, which suggests that a common Spanish population data-base could be possible. A semiparametric model is proposed for estimating frequencies, using the smoothed cross-validation of Hall et al. (1992) to calculate the size of the window utilized.

Alleles↗

PubMed1992

The use of immobilized pH gradients for the detection of human polymorphisms in the forensic identification of bloodstains.

Some polymorphic proteins (alpha 1-antitrypsin, orosomucoid, transferrin, group specific component, plasminogen) and enzymes (phosphoglucomutase, acid phosphatase, estrase D) were determined in bloodstain extracts by isoelectric focusing with carrier ampholytes (CA) and with immobilized pH gradients (IPGs) rehydrated with CA. IPGs yield superior results for typing of genetics markers in bloodstains since phenotypes are better distinguished and the bands are straighter and sharper in the presence of contaminants. Also, the sensitivity of IPGs with CA is similar to isoelectric focusing (IEF) with CA. A new variant, ACP*B1, found in Negroid west African populations and not found in Caucasians is described. Such a variant can only be determined by IPGs since its isoelectric point (pI 5.95) is close to that of the ACP*B (pI 6.05) variant.

Ampholyte Mixtures↗

PubMed1992

Detection of polymorphisms of human DNA after polymerase chain reaction by miniaturized SDS-PAGE.

PCR followed by SDS-PAGE in miniaturized non-denaturing gels permits in some cases the identification of single base pair substitutions in small DNA fragments and therefore, the study of human DNA polymorphisms. The usefulness of the system in forensic science is investigated by typing the HLA-DQA1 locus and the VNTR recognized with the probe pMCT118 (locus D1S80) and it shows to be advantageous over previously published methods for typing the MCT118 system, whereas in HLA-DQA1 typing for forensic casework, both dot-blot with ASO probes and this method could be complementary.

Base Sequence↗