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PubMed1998

Analysis of clonal diversity in mouse immunoglobulin heavy chain genes selected for size of the antigen combining site.

Size-diversity of Ig and T cell receptor antigen binding (CDR3) regions can be visualized by "CDR3 fingerprinting", and provides an estimate of B- or T-cell repertoire complexity. The method does not identify clonal diversity, however, which can only be determined by random sequencing of the CDR3s. In this study we demonstrate that a combination of fingerprinting and single strand conformation polymorphism (SSCP) analysis can be used for a rapid estimation of clonal diversity within mouse Ig antigen binding regions selected for size. This application may be useful in the analysis of clonal expansion within B- and T-cell repertoires.

Amino Acid Sequence↗
PubMed2004

[Analysis of genetic diversity of 14 species of Cymbidium based on RAPDs and AFLPs].

RAPDs and AFLPs were used to investigate the genetic variation of 14 species of Cymbidium. A great diversity was shown by 12 random primers and three primer combinations. UPGMA cluster analysis showed that the results from the RAPD and AFLP analysis were similar and the genetic distances were closest between C. ensifolium and C. sinense, C. kanran and C. faberi var. omeiense, C. mastersii and C. eburneum respectively.

Amplified Fragment Length Polymorphism Analysis↗
PubMed2002

Genomic diversity among Danish field strains of Mycoplasma hyosynoviae assessed by amplified fragment length polymorphism analysis.

Genomic diversity among strains of Mycoplasma hyosynoviae isolated in Denmark was assessed by using amplified fragment length polymorphism (AFLP) analysis. Ninety-six strains, obtained from different specimens and geographical locations during 30 years and the type strain of M. hyosynoviae S16(T) were concurrently examined for variance in BglII-MfeI and EcoRI-Csp6I-A AFLP markers. A total of 56 different genomic fingerprints having an overall similarity between 77 and 96% were detected. No correlation between AFLP variability and period of isolation or anatomical site of isolation could be demonstrated. An examination of the clonal appearance of M. hyosynoviae isolates recovered from seven herds affected with arthritis revealed presence of several genotypically distinct variants of the organism in a single herd, indicating that different strains may simultaneously be involved in an outbreak of the disease.

Animals↗
PubMed2004

Combined phospholipid biomarker-16S rRNA gene denaturing gradient gel electrophoresis analysis of bacterial diversity and physiological status in an intertidal microbial mat.

A combined lipid biomarker-16S rRNA gene denaturing gradient gel electrophoresis analysis was used to monitor changes in the physiological status, biomass, and microbial composition of a microbial mat. In the morning hours, an increase in the biomass of layers containing a high density of phototrophs and a decrease in the growth rate in the deep layers were observed. The combined approach also revealed differences in major groups of microorganisms, including green nonsulfur, gram-positive, and heterotrophic bacteria.

Bacteria↗
PubMed2003

Inter simple sequence repeat (ISSR) analysis of genetic diversity in tef [Eragrostis tef (Zucc.) Trotter].

The DNA polymorphism among 92 selected tef genotypes belonging to eight origin groups was assessed using eight inter simple sequence repeat (ISSR) primers. The objectives were to examine the possibility of using ISSR markers for unravelling genetic diversity in tef, and to assess the extent and pattern of genetic diversity in the test germplasm with respect to origin groups. The eight primers were able to separate or distinguish all of the 92 tef genotypes based on a total of 110 polymorphic bands among the test lines. The Jaccard similarity coefficient among the test genotypes ranged from 0.26 to 0.86, and at about 60 % similarity level the clustering of this matrix using the unweighted pair-group method based on arithmetic average (UPGMA) resulted in the formation of six major clusters of 2 to 37 lines with further eight lines remaining ungrouped. The standardized Nei genetic distance among the eight groups of origin ranged between 0.03 and 0.32. The UPGMA clustering using the standardized genetic distance matrix resulted in the identification of three clusters of the eight groups of origin with bootstrap values ranging from 56 to 97. The overall mean Shannon Weaver diversity index of the test lines was 0.73, indicating better resolution of genetic diversity in tef with ISSR markers than with phenotypic (morphological) traits used in previous studies. This can be attributed mainly to the larger number of loci generated for evaluation with ISSR analysis as compared to the few number of phenotypic traits amenable for assessment and which are further greatly affected by environment and genotype x environment interaction. Analysis of variance of mean Shannon Weaver diversity indices revealed substantial (P < or = 0.05) variation in the level of diversity among the eight groups of origin. In conclusion, our results indicate that ISSR can be useful as DNA-based molecular markers for studying genetic diversity and phylogenetic relationships, DNA fingerprinting for the identification of varieties or cultivars, and also for genome mapping in tef.

Electrophoresis, Polyacrylamide Gel↗
PubMed1991

[Diversity and relative inaccuracy of internal images of antigen group A revealed by IEF analysis].

The diversity of a polyclonal anti-idiotypic response (Ab2) to a murine monoclonal anti-A (Ab1) was investigated after purification of two Ab2 populations. One was eluted from human polyclonal anti-A column and the other from Ab1. Analysis of the Ab specificity, as well as screening of the clonotypic distribution, were achieved after splitting Ab by IEF; this was followed by immunoblotting and probing with various anti-ABH mAb. The first population reacted with almost all the murine anti-ABH mAb, as well as with four human anti-A mAb, and consequently consisted of Ab2 beta. The second was composed of "true" Ab2 directed against Ab1. In the first population internal images mimicked either A Ag, or H Ag, or some epitopes common to both. This study demonstrates the plurality of internal images-bearing Ig molecules, some mimicking completely, and some only partially or even unfaithfully the nominal A determinant. The analysis of this idiotypic cascade proves the existence of a degeneracy of the initial restricted antigenic specificity. The consequences of such a process are discussed.

Animals↗
PubMed2003

[Analysis of genetic diversity of Shandong indigenous chicken breeds using microsatellite marker].

Microsatellite marker is one of the frequently used molecular markers. It has been used in the genotype identification, pedigree analysis and estimation of genetic distance. In this paper, five microsatellite markers with high polymorphisms were selected to detect the genetic diversity of seven chicken breeds. The alleles frequencies, polymorphism information content (PIC) and average heterozygosity within each population, and DA genetic distance among breeds were analyzed. The application of microsatellite polymorphisms to the detection of genetic variability and relationship among populations was discussed. Altogether, forty alleles were found in this experiment, and among them the most alleles (10) were detected by ADL0136 and the least (5) were detected by ADL0146. The distribution of alleles was not balanced, each locus having one or more dominant alleles. The average heterozygosity in the Shouguang chicken was the lowest (0.3327), and that in other breeds was also less than 0.4. It can be seen then that microsatellite polymorphisms can be used to reveal the variability within population through calculation of average heterozygosity. The PIC values ranged from 0.6169 (Shouguang chicken) to 0.7027 (Laiwu Black chicken). UPGMA tree was completed through analysis of DA genetic distance. In the tree, the Rizhao Pockmarked and the Jining Hundred chicken were first grouped together with a bootstrap value of 92%, before they were grouped with the Laiwu Black and the Shouguang chickens. The Anoka Yellow and the Guangxi Yellow chicken were grouped together with a bootstrap value of 80%, but the Luxi Fighting chicken had its own branch. In summary, the UPGMA tree well reflected the evolutionary and breeding history of the seven breeds.

Alleles↗
PubMed2003

rep-PCR fingerptinting as a tool for the analysis of genomic diversity in Escherichia coli strains isolated from an aqueous/freshwater environment.

The rep-PCR fingerprinting method, with the support of ERIC and REP primers, was used to analyse the genomic diversity of 93 E. coli strains isolated from lake water samples drawn at two different depths. The applied UPGMA for DNA analysis did not reveale any genomic similarities between the 48 E. coli strains derived from the subsurface-zone water and the 43 of the bottom-zone water. The considerable genomic diversity of the E. coli of the surface zone was expressed as a dendrogram in the form of 8 similarity groups comprising strains isolated from samples drawn over one month. The bottom-zone strains, which display a lesser degree of genomic diversity (5 similarity groups), showed distinct common features in their DNA fingerprints. In the similarity dendrogram for the bottom-zone, strains derived in different months of sampling were segregated into the same similarity groups. Applying REP primers in rep-PCR generates more complex fingerprints increasing the discriminatory power of the analysis, whereas the ERIC primer generates less complex fingerprint patterns, and is thus clearer to interpret.

DNA Fingerprinting↗
PubMed1993

An analysis of the diversity of Haemophilus parainfluenzae in the adult human respiratory tract by genomic DNA fingerprinting.

A method for typing Haemophilus species is described, based on the analysis of genomic DNA from Haemophilus parainfluenzae. The DNA was extracted by a rapid method and digested with the restriction enzyme BamHI to provide a characteristic 'fingerprint'. The pattern of fragments in the ranges 1-1.6 kb, 1.6-2 kb and 2-3 kb were used to produce a numerical profile of each isolate. In total 97 isolates were examined; 88 from throat swab material isolated from the 15 members of a British Antarctic Survey base and 9 type strains. Seventy-two of the 88 antarctic isolates were H. parainfluenzae and were found to be very diverse, comprising 41 identifiable strains with up to 5 strains being isolated from a single throat swab sample. There was evidence for both carriage and transmission within the isolated community. The technique provided a highly discriminatory method for characterizing Haemophilus strains which is suitable for epidemiological studies.

Adult↗
PubMed2000

Mosaic plasmids and mosaic replicons: evolutionary lessons from the analysis of genetic diversity in IncFII-related replicons.

The alpha replicons of the multi-replicon plasmids pGSH500 and pLV1402 have been characterized by DNA sequence analysis. Analysis of the DNA sequence of a 3672 bp HIN:dIII fragment from pFDT100, which contains the pGSH500 alpha replicon, revealed similarity to a number of replicons belonging to, or related to, those of the IncFII family. The replicon region contains copA, tapA, repA and oriR, and replication initiation and termination sites are related to those from the IncFII replicon of R1. A copB gene was found to lie upstream of the HIN:dIII site in the parental plasmid pGSH500. Downstream of oriR, a 707 bp region shows 72.6% identity to a region of the Escherichia coli chromosome at 43.3', suggesting this region of pGSH500 may have been incorporated into the plasmid during a past chromosomal recombination event. Oligonucleotide primers homologous to consensus regions in the copB and repA genes, and the oriR regions from a number of IncFII-related replicons were used to amplify replication regions from pLV1402. Analysis of the amplified regions has shown the presence of copB, copA, tapA and repA genes. Phylogenetic analysis of Rep protein sequences from the RepFIIA family of antisense-control-regulated replicons revealed the presence of three distinct subgroups of Rep proteins. Comparative analysis of DNA and protein sequences from members of the RepFIIA family provides evidence supporting the roles of both non-selective divergence in co-integrate (multi-replicon) plasmids and Chi-mediated-recombination in replicon evolution, and in particular, that such processes may have been widespread in the evolution of the RepFIIA family.

Base Sequence↗
PubMed2005

Inter- and intraspecific resolution of nrDNA TRFLP assessed by computer-simulated restriction analysis of a diverse collection of ectomycorrhizal fungi.

An assessment of the inter- and intraspecific resolution of 5' nrDNA ITS TRFLP was conducted by computer-simulated restriction analysis of 316 ectomycorrhizal GenBank sequences. Generally, sequences with a similarity of < 90 % could be distinguished with two to three independent enzyme digests, although sequences with similarity > 95 % were likely to remain unresolved. Choice of restriction enzyme strongly influenced resolution, especially when less than three enzymes were used. Although our results showed that 5' nrDNA ITS TRFLP is a powerful tool for distinguishing between species of ectomycorrhizal fungi, closely related species, including species of Rhizopogon, Dermocybe, Hebeloma, and Lactarius produced indistinguishable 5' nrDNA ITS TRFLP--profiles. 5' nrDNA ITS TRFLP may also split a species, and the probability of this occurring reflected intraspecific sequence variation. For groups of closely related species, the use of 3' nrDNA ITS in conjunction with 5' nrDNA ITS produced improved resolution comparable with RFLP analyses. Overall, our results show that nrDNA ITS TRFLP is a valuable addition to the array of molecular tools available to ectomycorrhizal ecologists. However, simple assessments of ectomycorrhizal species diversity based on numbers of unique TRF created with a single restriction enzyme must be viewed with caution, as for each enzyme we examined, identical TRF common to groups of unrelated species were observed. In addition, to maximize the effectiveness of TRFLP in surveys of ectomycorrhizal fungi it will be useful to combine information from at least two independent enzyme digests in order to distinguish and track species in the field.

Basidiomycota↗
PubMed2004

Analysis of genetic diversity and conservation priorities in Iberian pigs based on microsatellite markers.

The Iberian breed is the most important pig population of the Mediterranean type. The genetic structure of two strains (Torbiscal and Guadyerbas) and three varieties (Retinto, Entrepelado and Lampiño) of this breed was studied using 173 pigs genotyped for 36 microsatellites. In addition, 40 pigs of the related Duroc breed were also analysed. In the 1960s, the Iberian breed's numbers were severely reduced by disease, due to economic change and to crossbreeding. Varieties are in danger of disappearance or blending. A new conservation strategy is required. An analysis was performed that allows us to ascertain the loss or gain of genetic diversity if one or several subpopulations are removed. The results are compared with those using the Weitzman method. The two methodologies produce conservation priorities that are completely different, the reason being that the Weitzman method does not take into account the within-population genetic diversity. We apply optimal contribution theory and a new procedure for cluster analysis, and discuss their value in the general framework of the problems of setting of priorities and tactics for the conservation of genetic resources.

Animals↗
PubMed2005

In vitro RNA random pools are not structurally diverse: a computational analysis.

In vitro selection of functional RNAs from large random sequence pools has led to the identification of many ligand-binding and catalytic RNAs. However, the structural diversity in random pools is not well understood. Such an understanding is a prerequisite for designing sequence pools to increase the probability of finding complex functional RNA by in vitro selection techniques. Toward this goal, we have generated by computer five random pools of RNA sequences of length up to 100 nt to mimic experiments and characterized the distribution of associated secondary structural motifs using sets of possible RNA tree structures derived from graph theory techniques. Our results show that such random pools heavily favor simple topological structures: For example, linear stem-loop and low-branching motifs are favored rather than complex structures with high-order junctions, as confirmed by known aptamers. Moreover, we quantify the rise of structural complexity with sequence length and report the dominant class of tree motifs (characterized by vertex number) for each pool. These analyses show not only that random pools do not lead to a uniform distribution of possible RNA secondary topologies; they point to avenues for designing pools with specific simple and complex structures in equal abundance in the goal of broadening the range of functional RNAs discovered by in vitro selection. Specifically, the optimal RNA sequence pool length to identify a structure with x stems is 20x.

Computational Biology↗
PubMed2006

Analysis of genetic diversity among Staphylococcus aureus isolates from patients with deep-seated and superficial staphylococcal infections using pulsed-field gel electrophoresis.

Molecular typing was performed to reveal the genetic diversity among Staphylococcus aureus strains causing deep-seated versus superficial staphylococcal infections. Pulsed-field gel electrophoresis with cluster analysis, plasmid and antimicrobial susceptibility profiling of 50 S. aureus strains collected from these 2 groups of patients were undertaken. A total of 19 (designated A through S) distinct genotypes were identified by PFGE of Sma I-digested genomic DNA. The most prevalent PFGE type was L, which accounted for 30% of isolates and was detected among superficial isolates only. The second most prevalent PFGE type, type A (18%), was predominant among deep-seated isolates. Remaining PFGE types varied in distribution between the 2 groups. Plasmid profile analysis revealed that deep isolates harbour plasmids more frequently (comprising 64% of isolates) than superficial isolates (4%) and showed 10 and 2 distinct patterns, respectively, with pattern 1 being the dominant among deep isolates. Antimicrobial susceptibility data suggested an increased prevalence of antibiotic resistance among deep isolates with the majority (40%) exhibiting identical antibiograms compared to superficial isolates. No resistance was detected against clindamycin and vancomycin. The results of our study indicate a previously unrecognized dichotomy of S. aureus strains, causing deep-seated and superficial infections.

Adolescent↗
PubMed2003

Genotypic diversity of oscillatoriacean strains belonging to the genera Geitlerinema and Spirulina determined by 16S rDNA restriction analysis.

Genotypic diversity of several cyanobacterial strains mostly isolated from marine or brackish waters, belonging to the genera Geitlerinema and Spirulina, was investigated by amplified 16S ribosomal DNA restriction analysis and compared with morphological features and response to salinity. Cluster analysis was performed on amplified 16S rDNA restriction profiles of these strains along with profiles obtained from sequence data of five Spirulina-like strains, including three representatives of the new genus Halospirulina. Our strains with tightly coiled trichomes from hypersaline waters could be assigned to the Halospirulina genus. Among the uncoiled strains, the two strains of hypersaline origin clustered together and were found to be distant from their counterparts of marine and freshwater habitat. Moreover, another cluster, formed by alkali-tolerant strains with tightly coiled trichomes, was well delineated.

Bacterial Proteins↗
PubMed2000

High resolution analysis of haplotype diversity and meiotic crossover in the human TAP2 recombination hotspot.

Little is known about the nature of recombination hotspots in the human genome and the relationship between crossover activity and patterns of linkage disequilibrium. We have therefore used both haplotype analysis and direct detection of crossovers in sperm to characterize a putative recombination hotspot in the TAP2 gene within the class II region of the MHC. Haplotype diversity provided evidence for a localized hotspot within intron 2 of this gene. Sperm DNA typing using allele-specific PCR primers to selectively amplify recombinant TAP2 molecules revealed a highly localized meiotic crossover hotspot approximately 1.2 kb long, unusually abundant in sequence polymorphisms and flanked by DNA much less active in recombination. Sperm crossover appeared to be fully reciprocal, and almost all crossover products were simple, involving a single exchange between adjacent heterozygous markers. This hotspot appears to be much more active in female than male meiosis. No primary sequence similarities could be found between any of the very few well defined crossover hotspots in the human genome, all of which show recombinationally active domains 1-2 kb long. Direct comparison of recombination frequency and haplotype diversity in TAP2 showed that linkage disequilibrium measures were a poor predictor of crossover frequency in this region, with non-recombining markers sometimes in free association and with examples of pairs of markers spanning the recombination hotspot showing substantial or even absolute linkage disequilibrium.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2006

AFLP analysis of genetic diversity of the endangered species Sinopodophyllum hexandrum in the Tibetan region of Sichuan Province, China.

Amplified fragment length polymorphism (AFLP) markers were used to estimate the genetic diversity of seven wild populations of Sinopodophyllum hexandrum (Royle) Ying from the Tibetan region of Sichuan Province, China. Six primer combinations generated a total of 428 discernible DNA fragments, of which 111 were polymorphic. The percentage of polymorphic bands (PPB) was 25.93 at the species level, and PPB within population ranged from 4.91 to 12.38%. Genetic diversity (H(E)) within populations varied from 0.01 to 0.04, averaging 0.05 at the species level. As revealed by the results of AMOVA analysis, 58.8% of the genetic differentiation occurred between populations, and 41.2% within populations. The genetic differentiation was, perhaps, due to the limited gene flow (Nm = 0.43) of the species. The correlation coefficient (r) between genetic and geographical distance using Mantel's test for all populations was 0.698 (P = 0.014). The UPGMA cluster analysis revealed a similar result in that the genetic distances among the populations show, to a certain extent, a spatial pattern corresponding to their geographic locations. On the basis of the genetic and ecological information, we propose some appropriate strategies for conserving the endangered S. hexandrum in this region.

Berberidaceae↗
PubMed2006

Analysis of genetic diversity and the determination of relationships among western Mediterranean horse breeds using microsatellite markers.

The distribution of genetic diversity and the genetic relationships among western Mediterranean horse breeds were investigated using microsatellite markers. The examined sample included seven Spanish and three Italian local horse breeds and populations, plus a Spanish Thoroughbred outgroup. The total number of animals examined was 682 (on average 62 animals per breed; range 20-122). The microsatellite marker set analysed provided 128 alleles (10.7 alleles per locus). Within-breed genetic diversity was always high (>0.70), with breeds contributing about 8% of the total genetic variability. The mean molecular coancestry of the entire population examined was 0.205, Losino being the breed that contributed most. In addition to Nei's standard and Reynolds' genetic distances, pair-wise kinship distance and molecular coancestry were estimated. Remarkably similar breed rankings were obtained with all methods. Clustering analysis provided an accurate representation of the current genetic relationships among the breeds. Determining coancestry is useful for analysing genetic diversity distribution between and within breeds and provides a good framework for jointly analysing molecular markers and pedigree information. An integrated analysis was undertaken to obtain information on the population dynamics in western Mediterranean native horse populations, and to better determine conservation priorities.

Alleles↗