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Various biomarkers currently used for the diagnosis of intestinal mucosal injury (IMI) in patients with acute intestinal obstruction have low sensitivity and specificity. In the present study, IMI, as indicated by the impaired expression of tight junction proteins, including zonula occludens-1, occludin and claudin-1, and inflammation were determined in colonic tissues of patients with 45 strangulated intestinal obstruction (STR-IO) and the adjacent "normal" colonic tissues of 35 patients with colon cancers by quantitative real-time polymerase chain reaction (QRT-PCR), Western blotting, immunohistochemistry and histological examination, respectively. Then, two-dimensional fluorescent difference gel electrophoresis coupled with linear trap quadrupole mass spectrometry was used to screen for potential biomarkers of IMI in the serum samples of 10 STR-IO, 10 simple intestinal obstruction (SIM-IO) and 10 normal healthy controls. A total of 35 protein spots were differentially expressed among the serum samples, and six of the proteins were identified as potential biomarkers. Among the six proteins, histidine decarboxylase (HDC) and ceruloplasmin (CP) were elevated significantly in patients with STR-IO, compared with patients with SIM-IO and healthy controls. Thus, HDC and CP were further validated by QRT-PCR, Western blotting, immunohistochemistry and enzyme-linked immunosorbent assay, respectively, in colonic tissues, serum and urine samples. Finally, the receiver operating characteristic curves were used to show the area under the curves of HDC, CP and several established biomarkers, followed by the determination of the appropriate cutoff values and their sensitivities and specificities. It was shown that for serum and urine, HDC levels achieved sensitivities and specificities compatible to or even greater than those of established biomarkers for the diagnosis of IMI in patients with acute intestinal obstruction, although further validation in a larger cohort is required.
Mol Med 2011
PMID:Histidine decarboxylase is identified as a potential biomarker of intestinal mucosal injury in patients with acute intestinal obstruction. 2191 37

Allele frequencies of ten autosomal STR loci, D3S1358, vWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, TH01, and FGA were investigated in Chakma and Tripura tribal populations of Bangladesh. In both the populations, all loci were in Hardy-Weinberg equilibrium except for FGA locus in Chakma and D21S11 in Tripura. All the loci were highly polymorphic in Chakma population with an observed heterozygosity (Ho) of >0.7 and moderately polymorphic in Tripura population (Ho > 0.6). However, both the population showed least polymorphism at TH01 locus (Ho < 0.6). A comparison between Chakma and Tripura population data revealed statistically significant differences in allele frequency distribution for most of the loci. A similar comparison with the mainstream Bengali population using previously published data from this lab also showed significant difference in allele frequency with these two tribal populations.
Mol Biol Int 2010
PMID:Allele Frequencies of 10 Autosomal STR Loci from Chakma and Tripura Tribal Populations in Bangladesh. 2211 Sep 59

This chapter presents a brief introduction to the historical development of current technologies used in DNA analysis for human identification. The text describes the development of the PCR and short tandem repeats along with subsequent advances in instrumentation such as real-time PCR and capillary electrophoresis. These techniques have brought about a revolution in DNA typing methods through increased efficiency and the application of multiplex fluorescence detection. More recently the development of new STR based typing methods utilizing mini- and Y-STR PCR multiplexes has increased the flexibility of the investigator, permitting the analysis of inhibited and degraded DNA. Future directions for DNA typing are also discussed, including the development of methods for touch samples based on low copy DNA analysis and the determination of tissue/cell type.
Methods Mol Biol 2012
PMID:An overview of DNA typing methods for human identification: past, present, and future. 2213 49

Y-STR analysis is an invaluable tool to examine evidence in sexual assault cases and in other forensic casework. Unambiguous detection of the male component in DNA mixtures with a high female background is still the main field of application of forensic Y-STR haplotyping. In the last years, powerful technologies including a 17-locus multiplex PCR assay have been introduced in the forensic laboratories. At the same time, statistical methods have been developed and adapted for interpretation of a nonrecombining, linear marker as the Y-chromosome which shows a strongly clustered geographical distribution due to the linear inheritance and the patrilocality of ancestral groups. Large population databases, namely the Y-STR Haplotype Reference Database (YHRD), have been established to assess the evidentiary value of Y-STR matches by means of frequency estimation methods (counting and extrapolation).
Methods Mol Biol 2012
PMID:Interpretation guidelines of a standard Y-chromosome STR 17-plex PCR-CE assay for crime casework. 2213 52

During the two last decades, STR markers located on the autosomes have been gaining relevance and have nearly replaced the use of other type of markers in most cases of genetic identification, paternity testing, as well as in other situations of kinship analysis. Nevertheless, in some complex cases, independently of the number of polymorphisms being typed, autosomal markers convey very little information. Depending on the parentage constellation available for analysis, as well as the gender of the subjects, this problem can sometimes be solved by using markers that have different modes of transmission. Therefore, most forensic laboratories are nowadays prepared to analyse lineage markers (Y chromosome and mtDNA) and many have recently set up methods for the analysis of X-STRs. In the present chapter, a method is described for the typing of ten X chromosome-specific markers in a single PCR amplification reaction, followed by capillary electrophoresis separation and fluorescent detection in an ABI Genetic Analyser apparatus. This typing strategy was developed and optimized for the simultaneous amplification of ten X-linked specific STRs well distributed along the chromosome: DXS8378, DXS9902, DXS7132, DXS9898, DXS6809, DXS6789, DXS7133, GATA172D05, GATA31E08 and DXS7423.
Methods Mol Biol 2012
PMID:Capillary electrophoresis of an X-chromosome STR decaplex for kinship deficiency cases. 2213 53

Ancestry inference based on autosomal markers remains a niche approach in forensic analysis: most laboratories feel more secure with a review of the cumulative STR profile frequencies in a range of relevant populations with the possible additional analysis of mitochondrial and/or Y-chromosome variability. However, a proportion of autosomal single nucleotide polymorphisms (SNPs) show very well-differentiated allele frequencies among global population-groups. Furthermore, such ancestry informative marker SNPs (AIM-SNPs) lend themselves to relatively straightforward typing with short-amplicon PCR and multiplexed single base extension reactions using the same capillary electrophoresis detectors required for the sequencing and STR genotyping of mainstream forensic markers. In this chapter, we describe a 34 AIM-SNP multiplex that is robust enough for the analysis of challenging, often highly degraded DNA typical of much of routine forensic casework. We also outline in detail the in-silico procedures necessary for collecting parental population reference data from the SPSmart SNP databases and performing ancestry inference of single AIM-SNP profiles or large-scale population data using the companion ancestry analysis website of Snipper. Two casework examples are described that show, in both cases, that an inference of likely ancestry using AIM-SNPs helped the identification of highly degraded skeletal material.
Methods Mol Biol 2012
PMID:A 34-plex autosomal SNP single base extension assay for ancestry investigations. 2213 56

This work describes the main advantages and the steps involved in the optimization of a multiplex system able to characterize 38 noncoding biallelic Insertion Deletion Polymorphisms(Indels). With this methodology, all markers are amplified in a single PCR, using short amplicons (up to 160 bp) in order to improve its performance in degraded samples. Alleles are easily detected using capillary electrophoresis.The Indel multiplex typing strategy here described has the same desirable characteristics of forensic SNP assays, including genetic markers (a) with low mutation rates, increasing their usefulness in some kinship cases where few or single incompatibilities can be explained by mutation, and (b) that can be typed using a short amplicon strategy, increasing their usefulness in cases where degraded samples are available. Moreover, this approach uses simple and well-established methodologies already applied in forensic STR assays.
Methods Mol Biol 2012
PMID:Capillary electrophoresis of 38 noncoding biallelic mini-Indels for degraded samples and as complementary tool in paternity testing. 2213 58

STR analysis of DNA extracted from skeletal samples can play an important role in the identification of missing persons. Here we present a method for the extraction of DNA from skeletal samples involving complete demineralization and digestion of the sample, followed by purification by silica binding. This method, together with the multiplex STR typing approach also presented, has proven highly successful in the recovery of DNA profiles from degraded, aged skeletal remains from a wide range of environmental contexts. The methodological steps presented include bone decontamination and grinding, DNA extraction, repurification in the case of highly inhibited samples, quantification, STR multiplex amplification, and profile reporting guidelines. However, the conditions applied for amplification and the criteria for allele calling and profile submission must be based on the results of each laboratory's internal validation experiments involving the type of samples relevant to the project at hand. The methods presented here have permitted large-scale DNA-based identification of persons missing from mass disasters and armed conflict.
Methods Mol Biol 2012
PMID:DNA extraction from aged skeletal samples for STR typing by capillary electrophoresis. 2213 61

The interpretation of multilocus STR profiling is a complex task that requires a high degree of expertise due to the high number of variables that can affect a biological and analytical process like this.The purpose of this chapter is to provide guidelines to categorize the mixtures using a discrete system and select those in the white area to produce a profile for reporting and/or comparing.
Methods Mol Biol 2012
PMID:Interpretation guidelines for mixed-STR multilocus electrophoretic profiles. 2213 63

STR analysis of canine-derived biological evidence for the identification of individuals is becoming an important tool for forensic investigations. A protocol for the multiplex PCR amplification and capillary electrophoresis of nine autosomal STRs and two fixed-size markers for sex identification in dogs and wolves is described here. The selection of the loci included in the multiplex complies with the recommendations of the International Society for Forensic Genetics in regard to human DNA analysis. The protocol is optimized for automatic fragment size detection in an ABI platform.
Methods Mol Biol 2012
PMID:Capillary electrophoresis analysis of a 9-plex STR assay for canine genotyping. 2213 64


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