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PubMed1993

The chromosomal location of genes for elongation factor Tu and ribosomal protein S10 in the cyanobacterium Spirulina platensis provides clues to the ancestral organization of the str and S10 operons in prokaryotes.

The structural gene (rps10) encoding ribosomal protein S10 of the cyanobacterium Spirulina platensis has been localized both on chromosomal DNA and the previously characterized recombinant plasmid pSp7 harbouring the 3'-terminal portion of the gene for elongation factor G (fus) and the gene for elongation factor Tu (tuf). Alignment of the predicted S10 sequence of S. platensis with the homologous sequences from cyanelles, bacteria, archaea and eukarya showed that the cyanobacterial S10 shares a high degree of sequence homology (74% amino acid identity) with the cyanellar protein. Unlike the situation in Escherichia coli, the rps10 gene of S. plantensis is unlinked to the S10 operon genes, being adjacent to the str operon genes. Since a similar organization could be observed in cyanelles of Cyanophora paradoxa and in all archaea so far analysed, this probably represents the ancestral state.

Amino Acid Sequence↗
PubMed1993

Nucleotide sequence of the first cosmid from the Mycobacterium leprae genome project: structure and function of the Rif-Str regions.

The nucleotide sequence of cosmid B1790, carrying the Rif-Str regions of the Mycobacterium leprae chromosome, has been determined. Twelve open reading frames were identified in the 36716bp sequence, representing 40% of the coding capacity. Five ribosomal proteins, two elongation factors and the beta and beta' subunits of RNA polymerase have been characterized and two novel genes were found. One of these encodes a member of the so-called ABC family of ATP-binding proteins while the other appears to encode an enzyme involved in repairing genomic lesions caused by free radicals. This finding may well be significant as M. leprae, an intracellular pathogen, lives within macrophages.

Bacterial Proteins↗
PubMed2006

Incestuous paternity detected by STR-typing of chorionic villi isolated from archival formalin-fixed paraffin-embedded abortion material using laser microdissection.

Microscopic examination of a blood clot expelled by a physically and mentally disabled woman taken to the emergency room because of genital bleeding revealed the presence of chorionic villi encircled by decidua, hemorrhage, and necrosis. In order to identify the father of the product of conception, sections of formalin-fixed, paraffin-embedded abortion material were subjected to laser microdissection: DNA extraction from chorionic villi selectively isolated from the surrounding tissues allowed successful STR-typing of fetal cells, which was otherwise prevented by excess maternal DNA. The large number of homozygous genotypes in the fetal profile suggested incestuous paternity. Analysis of reference DNA samples from male relatives excluded the woman's father, paternal grandfather, and maternal grandfather, whereas the obligate paternal alleles of the fetus were constantly present in the genotypes of the woman's brother, clearly demonstrating brother-sister incest (probability of paternity > 99.99999%).

Aborted Fetus↗
PubMed2006

A proposal for standardization in forensic canine DNA typing: allele nomenclature of six canine-specific STR loci.

In this study a proposal for the allele nomenclature of six polymorphic short tandem repeat (STR) loci (PEZ3, PEZ6, PEZ8, PEZ10, FHC2161, and FHC2328) for canine genotyping (Canis lupus familiaris) is presented. The nomenclature is based on the sequence data of the polymorphic region of the microsatellite markers as recommended by the DNA commission of the International Society of Forensic Haemogenetics (ISFH) in 1994 for human DNA typing. To cover commonly and rarely occurring alleles, a selection of homozygous and heterozygous animals were analyzed and subjected to sequence studies. The alleles consisted of simple tri- and tetra-nucleotide repeat patterns as well as compound and highly complex repeat patterns. Several alleles revealing the same fragment size but different repeat structures were found. The allele designation described here was adopted to the number of repeats, including all variable regions within the amplified fragment. In a second step the most commonly occurring alleles were added to an allelic ladder for each marker allowing a reliable typing of all alleles differing in size. A total number of 142 unrelated dogs from surrounding municipal animal homes, private households, and canines in police duty were analyzed. The data were added to a population database providing allele frequencies for each marker.

Alleles↗
PubMed2007

Run-specific limits of detection and quantitation for STR-based DNA testing.

STR-based DNA profiling is an exceptionally sensitive analytical technique that is often used to obtain results at the very limits of its sensitivity. The challenge of reliably distinguishing between signal and noise in such situations is one that has been rigorously addressed in numerous other analytical disciplines. However, an inability to determine accurately the height of electropherogram baselines has caused forensic DNA profiling laboratories to utilize alternative approaches. Minimum thresholds established during laboratory validation studies have become the de facto standard for distinguishing between reliable signal and noise/technical artifacts. These minimum peak height thresholds generally fail to consider variability in the sensitivity of instruments, reagents, and the skill of human analysts involved in the DNA profiling process over the course of time. Software (BatchExtract) made publicly available by the National Center for Biotechnology Information now provides an alternative means of establishing limits of detection and quantitation that is more consistent with those used in other analytical disciplines. We have used that software to determine the height of each data collection point for each dye along a control sample's electropherogram trace. These values were then used to determine a limit of detection (the average amount of background noise plus three standard deviations) and a limit of quantitation (the average amount of background noise plus 10 standard deviations) for each control sample. Analyses of the electropherogram data associated with the positive, negative, and reagent blank controls included in 50 different capillary electrophoresis runs validate that this approach could be used to determine run-specific thresholds objectively for use in forensic DNA casework.

DNA Fingerprinting↗
PubMed1982

Evidence for an internal promoter preceding tufA in the str operon of Escherichia coli.

We constructed plasmids carrying tufA from which the major promoter for the rpsL-rpsG-fus-tufA operon (also called the str operon) had been removed. These plasmids continued to express tufA, as judged by the ability to complement mocimycin resistance and by electrophoretic analysis of synthesized proteins. Tn5 transpositions into fus, the gene for elongation factor G, which lies immediately on the 5' side of tufA, failed to obstruct the expression of tufA. The subcloning of a 2,000-base-pair PstI-SmaI DNA fragment (containing the intercistronic region between tufA and fus, the distal portion of fus, and the proximal portion of tufA) next to promoterless tetracycline resistance genes (tet) yielded a plasmid that was capable of bestowing resistance to 12 microgram of tetracycline per ml. The removal of an EcoRI fragment that lies within fus destroyed the ability of the 2,000-base-pair PstI-SmaI fragment to promote the transcription of tet. These data indicate that, in addition to the operon's major promoter rpsLp, there is an internal promoter, tufAp, which can be used for the transcription of tufA, tufAp probably lies within fus, about 50 base pairs upstream from its 3' end and 120 base pairs from the start codon of tufA. The relative activities of tufB and of tufA-from-tufAp were estimated by a comparison of beta-galactosidase activities of almost identical EF-Tu-beta-galactosidase protein fusions; they were approximately equal.

Cloning, Molecular↗
PubMed1985

Self-cloning in Streptomyces griseus of an str gene cluster for streptomycin biosynthesis and streptomycin resistance.

An str gene cluster containing at least four genes (strR, strA, strB, and strC) involved in streptomycin biosynthesis or streptomycin resistance or both was self-cloned in Streptomyces griseus by using plasmid pOA154. The strA gene was verified to encode streptomycin 6-phosphotransferase, a streptomycin resistance factor in S. griseus, by examining the gene product expressed in Escherichia coli. The other three genes were determined by complementation tests with streptomycin-nonproducing mutants whose biochemical lesions were clearly identified. strR complemented streptomycin-sensitive mutant SM196 which exhibited impaired activity of both streptomycin 6-phosphotransferase and amidinotransferase (one of the streptomycin biosynthetic enzymes) due to a regulatory mutation; strB complemented strain SD141, which was specifically deficient in amidinotransferase; and strC complemented strain SD245, which was deficient in linkage between streptidine 6-phosphate and dihydrostreptose. By deletion analysis of plasmids with appropriate restriction endonucleases, the order of the four genes was determined to be strR-strA-strB-strC. Transformation of S. griseus with plasmids carrying both strR and strB genes enhanced amidinotransferase activity in the transformed cells. Based on the gene dosage effect and the biological characteristics of the mutants complemented by strR and strB, it was concluded that strB encodes amidinotransferase and strR encodes a positive effector required for the full expression of strA and strB genes. Furthermore, it was found that amplification of a specific 0.7-kilobase region of the cloned DNA on a plasmid inhibited streptomycin biosynthesis of the transformants. This DNA region might contain a regulatory apparatus that participates in the control of streptomycin biosynthesis.

Amidinotransferases↗
PubMed1985

Serological identification of Streptococcus sanguis and Str mitior.

A total of 165 strains of Streptococcus sanguis and Str mitior were selected on the basis of their biochemical reactions using established identification procedures. These strains were also classified using API Database and were then screened against five candidate grouping sera. Biochemical tests and serological identification were in general complementary, but no regular associations between biotype and serological reaction were observed.

Serotyping↗
PubMedDate not supplied

Allelic affinities between the F13A common gene products inferred by the analysis of an (AAAG)n STR polymorphism within the 5' untranslated region.

Factor XIII a subunit (F13A) is the last enzyme in the blood coagulation cascade. It is characterized by extensive genetic polymorphism defined by 4 common alleles, F13A*1A, 1B, 2A and 2B and a few rare variants, some responsible for severe coagulation deficiencies. In order to infer the evolutionary affinities between the common F13A alleles we have applied PCR techniques to study, in a Northern Portuguese sample, a short tandem repeat polymorphism located within the 5' untranslated region of the F13A gene. The analysis of the molecular heterogeneity within the F13A gene products revealed that the four biochemical variants shared very similar, truncated, distributions of STR alleles and showed no signs of predominant haplotypic associations. These findings seem to support both the inferences that intragenic recombination played an important role in the generation of molecular diversity within each of the four main F13A alleles and that all the four F13A alleles must be rather old. Molecular heterogeneity levels allowed the identification of 1B as the oldest F13A allelic state, and 2A as the most recently generated allele, but were not different enough to accurately track the divergence of alleles 1A and 2B. However, additional analysis of linkage disequilibrium patterns indicates that 1B-->2B-->1A-->2A is the most likely evolutionary order of appearance of F13A main protein alleles, confirming and extending a previous hypothetical model inferred from their molecular features.

5' Untranslated Regions↗
PubMed2004

Analysis of the cytogenetic stability of the human embryonal kidney cell line 293 by cytogenetic and STR profiling approaches.

We have characterized the cytogenetic alterations of the human embryonal cell line 293 by spectral karyotyping and G-banding analysis. To investigate its genomic stability, we compared the karyotypes of 293 and its daughter line EcR-293. Genotype profiling through short tandem repeats complemented the analysis. While displaying almost identical STR profiles and thus verifying their origin and their close relation, the two lines were remarkably different in their number of chromosomes and setup of aberrant chromosomes. However, the cell lines retained a stable karyotype in long term culture. The establishment of subclones from EcR-293, expressing inducible lacZ or MEN1 transgenes, only added minor changes to the karyotype. Our study shows that the cytogenetic constitution of a clonal cell line of the 293 origin appears to be sufficiently stable. However, care should be taken when comparing the properties of independent 293 lineages, since clonal variations might be substantial.

Cell Culture Techniques↗
PubMed1991

The nucleotide sequences of Bacillus stearothermophilus ribosomal protein S12 and S7 genes: comparison with the str operon of Escherichia coli.

The primary structure of the ribosomal protein S7 from Bacillus stearothermophilus (BstS7) was analyzed by a combination of amino acid and DNA sequence analysis. Peptide sequence information was derived from tryptic peptides of BstS7 by manual Edman degradation. The nucleotide sequence of the 1.5-kb SalI fragment from B. stearothermophilus genome, cloned by the Escherichia coli S12 gene (rpsL) as a hybridization probe, was determined. Comparison of deduced amino acid sequence with the corresponding sequences of E. coli ribosomal proteins showed that this fragment contains the genes encoding S12, S7, and the N-terminus of the elongation factor G. Thus, the organization of this gene cluster is same as that in the str operon of E. coli. The amino acid sequence of B. stearothermophilus S12 (BstS12) deduced from the nucleotide sequence information agrees with the published amino acid sequence of BstS12 [M. Kimura and J. Kimura, FEBS Lett., 210, 91 (1987)]. The deduced sequence of B. stearothermophilus S7 (BstS7), together with sequence information of tryptic peptides, showed that protein S7 consists of 155 amino acid residues with a calculated molecular weight of 17933 and has 56% amino acid identity with the E. coli S7 (EcoS7).

Amino Acid Sequence↗
PubMed2001

Variability of six STR loci in populations from Central Spain.

Allele frequency distributions of six short tandem repeat (STR) loci, HUMTH01, HUMFES/FPS, HUMTPOX, HUMVWF/A31, HUMF13B, and HUMLPL, were determined in a population (101 individuals) of the Vera-Jerte region in West Central Spain. Amplified products were electrophoresed on denaturing polyacrylamide gels and silver-stained. The exact test demonstrated that none of the six loci deviated from Hardy-Weinberg equilibrium. R-matrix analysis in relation to other European samples agrees well with genetic distance results of Reynolds et al. (1983). Comparisons show that our population comes reasonably well within the range of variation of other European samples, and that the representation of these samples on the genetic map indicates a close relation between geographical location and distribution in the diagram.

Alleles↗
PubMed2004

Minimal sharing of Y-chromosome STR haplotypes among five endogamous population groups from western and southwestern India.

We attempt to address the issue of genetic variation and the pattern of male gene flow among and between five Indian population groups of two different geographic and linguistic affiliations using Y-chromosome markers. We studied 221 males at three Y-chromosome biallelic loci and 184 males for the five Y-chromosome STRs. We observed 111 Y-chromosome STR haplotypes. An analysis of molecular variance (AMOVA) based on Y-chromosome STRs showed that the variation observed between the population groups belonging to two major regions (western and southwestern India) was 0.17%, which was significantly lower than the level of genetic variance among the five populations (0.59%) considered as a single group. Combined haplotype analysis of the five STRs and the biallelic locus 92R7 revealed minimal sharing of haplotypes among these five ethnic groups, irrespective of the similar origin of the linguistic and geographic affiliations; this minimal sharing indicates restricted male gene flow. As a consequence, most of the haplotypes were population specific. Network analysis showed that the haplotypes, which were shared between the populations, seem to have originated from different mutational pathways at different loci. Biallelic markers showed that all five ethnic groups have a similar ancestral origin despite their geographic and linguistic diversity.

Chromosomes, Human, Y↗
PubMed2005

Characterization of the D18S535 STR locus in northern Ontario European and Aboriginal populations for forensic purposes.

Genetic characterization of one European and three aboriginal populations from northern Ontario was undertaken to assess the utility of the D18S535 short tandem repeat locus (STR) as a genetic marker for forensic DNA typing in the region. The D18S535 locus was amplified using monoplex polymerase chain reaction (PCR), separated by denaturing polyacrylamide gel electrophoresis (PAGE), and visualized using the silver-stain detection method. The generated population data demonstrated that the D18S535 locus is highly polymorphic with a heterozygosity of > or = 0.75. The exact test showed violations of Hardy-Weinberg equilibrium in two of the aboriginal populations. Pairwise comparisons of allele-frequency distributions indicated that the four northern Ontario populations were significantly different from each other. This test also revealed that the northern Ontario populations differed significantly from ten European populations (from Germany, Spain, and Croatia) and one population from South America (from Argentina). Forensic parameters showed that the D18S535 locus is highly discriminating (power of discrimination > or = 0.85, chance of exclusion > or = 0.51); however, the lack of Hardy-Weinberg equilibrium in some of the populations must be taken into account in the application of these results to northern Ontario forensic casework.

American Indian or Alaska Native↗
PubMed2006

STR markers for kinship analysis.

The analysis of short tandem repeats is a widely used method to estimate relatedness between closely related populations or individuals. The AmpFlSTR PCR Amplification Kit has 15 highly variable autosomal markers of tetranucleotide repeats and is principally made to identify individuals and first- or second-degree relatives. However, in many studies one is searching for individuals who are related through more than one generation. We wanted to test whether the amplification kit can also be used to identify more distantly related individuals. Therefore we compared 16 different methods that calculate genetic distance with regard to each method's ability to cluster more distantly related individuals from two test families. Among all the tested methods Nei et al.'s (1983) DA distance performed well in clustering family members within a group of unrelated individuals for a broad range of scenarios. However, second-degree relatives were difficult to cluster with any of the examined methods when other family members were absent. With a simulation we further estimated how many markers would actually be needed to detect a certain degree of relatedness. According to this simulation, one would need at least 123 independent microsatellite markers to detect third-degree relatives with 90% probability. In conclusion, the 15 STR markers in the amplification kit are suitable for detecting only very closely related individuals or entire families.

Female↗
PubMed2005

Y chromosome STR typing.

Because of their unique transmission properties and male specificity, markers located on the nonrecombining region of the Y chromosome (NRY) have become an important tool in forensic investigation. In the past few years, more than 50 polymorphic Y chromosome-specific short tandem repeats (STRs) have been described and a set of 9 loci were selected, considered as the minimal haplotype included in the forensic databases, Y-STR Haplotype Reference Databases, the largest now available online (http://www.yhrd.org). Here, we describe a multiplex amplification strategy developed for ABI platforms to amplify the nine loci included in the minimal haplotype (DYS19, DYS385a/b, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393) as well as a second set of markers (DYS437, DYS438, DYS439, DYS460, DYS461, GATA A10, GATA C4, and GATA H4) used to improve the minimal haplotype diversity and, hence, the discrimination power. Together, these 17 Y-STRs can be used to define highly discriminative haplotypes, providing a powerful tool in male lineage identification.

Base Sequence↗
PubMed1997

Polymorphism of short tandem repeat (STR) loci PLA2, D3S2459, D8S315 (kw38), CYP19, D3S1359, FGA, and D8S1132 in the Japanese and Chinese populations.

We report a study of polymorphism for seven short tandem repeat (STR) loci in Japanese and Chinese populations. Among 104 to 134 individuals in the both population samples, eight alleles were revealed for locus PLA2, thirteen for D3S1359, eleven for FGA, eight for D8S315 (kw38), ten for D8S1132, five for CYP19, and seven for D3S2459. They correspondingly constituted 10 to 39 genotypes therein. For most of the STRs, there was only a single allele active as the most frequent one among the others, except locus D3S1359 in Chinese samples (two alleles, 206 bp and 210 bp, frequency = 0.273 each). Also, the population genotype configurations were locus specific, varying in the patterns of commonest genotypes on each locus, e.g., one pattern for loci CYP19, D3S1359, and D8S315, one and two for loci PLA2 and D3S2459, two for locus D8S1132, and one and four for locus FGA. The distributions of observed genotypes were in Hardy-Weinberg Equilibrium. Furthermore, the seven STRs were exhibited highly polymorphic and informative for the both populations, and the alleles could be easily separated in electrophoresis and correctly interpreted with side-to-side allelic ladders. Together, the results suggest that the tri- and tetra-meric STRs are useful genetic markers for forensic practice.

Alleles↗
PubMed2001

Fitness of the marine parasitic nematode Anisakis simplex s. str. in temperate waters of the NE Atlantic.

In temperate waters of the NE Atlantic, third-stage larvae of Anisakis simplex sensu stricto collected from 3 paratenic host species were identified by restriction fragment length polymorphisms. The condition of wild larval infrapopulations was assessed by examining morphometric and growth characteristics. The differentiation patterns and the excretory/secretory products of larvae grown in vitro were also examined. An extensive morphometric, growth and differentiation variability was found between parasite larvae collected from different paratenic host sources. Nematode infrapopulation larvae from the squid comprise those smaller individuals with the lowest values of survival rates and moult success. It may be then concluded that the fitness of A. simplex s. str. larvae is not the best possible in the squid, which impaired the competitiveness of the parasite and its chances of developing into an adult. This suggests that the microenvironments impaired by the paratenic host may provide larval infrapopulations with unique ecological factors probably influencing its recruitment to the final host populations.

Animals↗