Individual Identification and Paternity Determination in Chimpanzees (Pan troglodytes) Using Human Short Tandem Repeat (STR) Markers

We studied 155 human short tandem repeat (STR) DNA markers in chimpanzees (Pan troglodytes) via the polymerase chain reaction (PCR). There is no difference in number of alleles per locus among STRs of different motif length (di-, tri-, or tetranucleotide repeats). We investigated 42 of the most informative STRs in greater detail using DNA isolated from a panel of 41 African-born, captive-housed chimpanzees. They reveal a wealth of genetic variability in chimpanzees, with an average of six alleles and 70.6% heterozygosity. The average paternity exclusion probability is 51.6%, and the best three STRs jointly provide >95% mean exclusion probability. Used in combination to define a multiple-locus genotype, the five most informative focal STRs can potentially uniquely identify every chimpanzee alive in the world. Although the subjects are of unknown geographical origin, homozygosity tests indicate little evidence for population subdivision. These markers represent the basis of a powerful battery of genetic tests, including individual identification, e.g., in poaching, paternity testing, or reconstruction of pedigrees among captive and wild chimpanzee breeding populations.     

Similarity of Structural Features and Evolution of Satellite DNAs from Palorus subdepressus (Coleoptera) and Related Species

A novel highly abundant satellite DNA comprising 20% of the genome has been characterized in Palorus subdepressus (Insecta, Coleoptera). The 72-bp-long monomer sequence is composed of two copies of T₂A₅T octanucleotide alternating with 22-nucleotide-long elements of an inverted repeat. Phylogenetic analysis revealed clustering of monomer sequence variants into two clades. Two types of variants are prevalently organized in an alternating pattern, thus showing a tendency to generate a new complex repeating unit 144 bp in length. Fluorescent in situ hybridization revealed even distribution of the satellite in the region of pericentric heterochromatin of all 20 chromosomes. P. subdepressus satellite sequence is clearly species specific, lacking similarity even with the satellite from congeneric species P. ratzeburgii. However, on the basis of similarity in predicted tertiary structure induced by intrinsic DNA curvature and in repeat length, P. subdepressus satellite can be classified into the same group with satellites from related tenebrionid species P. ratzeburgii, Tenebrio molitor, and T. obscurus. It can be reasonably inferred that repetitive sequences of different origin evolve under constraints to adopt and conserve particular features. Obtained results suggest that the higher-order structure and repeat length, but not the nucleotide sequence itself, are maintained through evolution of these species. Received: 23 April 1997 / Accepted: 11 July 1997     

Population database of STRs in West Africa: a genetic study of TPOX, HUMVWA31/A, HUMTH01, and CYP19

Four tetrameric STRs (TPOX, HUMVWA31/A, HUMTH01, and CYP19) were analysed in a West African population (Cabo Verde). No significant deviations from Hardy–Weinberg proportions were observed, either in conventional or exact tests. Pairwise comparisons confirmed allelic independence for all the combinations of loci. Data is provided for the first time about CYP19 in Black populations. In comparisons between African and Afro‐American populations, significant frequency differences for several alleles at the TH01 and VWA31/A loci were observed. The allele frequencies provided in this study contribute to a better knowledge of the variability of these markers among the main human groups, especially in the context of Subsaharan African populations. This revised version was published online in July 2006 with corrections to the Cover Date.     

Length polymorphism and allele structure of trinucleotide microsatellites in natural accessions of Arabidopsis thaliana

 The objective of this work was to assess the degree of trinucleotide microsatellite length polymorphism in the selfing species Arabidopsis thaliana. PCR amplifications of 12 microsatellite loci among 49 natural populations revealed between one to eight length variants (alleles) for each locus. The average number of alleles per locus was four and the average genetic diversity index was 0.43. Divergence between length variants was investigated at the nucleotide level. Several observations emerge from the sequence data: (1) for most loci, length polymorphism results only from variations in the number of trinucleotide repeats; (2) for a few others, some variability was noted in the flanking sequences; (3) for compound and interrupted loci containing two arrays of trinucleotide repeats, length variations preferentially affect the longest one. Five of the Arabidopsis thaliana accessions were clearly composed of two sublines. In 2 other accessions, some heterozygous individual plants, probably resulting from recent outcrosses, were found. A phylogenetic tree constructed on the basis of trinucleotide microsatellite allelic diversity shows that genetic relationships among the accessions are not correlated with their geographic origin. Received: 4 November 1997 / Accepted: 3 March 1998     

Rpg1, a soybean gene effective against races of bacterial blight, maps to a cluster of previously identified disease resistance genes

 Alleles, or tightly linked genes, at the soybean (Glycine max L. Merr.) Rpg1 locus confer resistance to races of Pseudomonas syringae pv. glycinea that express the avirulence genes avrB or avrRpm1. In this study we demonstrate that Rpg1 maps to a cluster of previously identified resistance genes, including those effective against fungal, viral and nematode pathogens. Rpg1 is in molecular linkage group (MLG) F, flanked by the markers K644 and B212. The RFLP markers R45, php2265 and php2385 cosegregated with Rpg1, as did the marker nbs61, which encodes a protein related to previously isolated resistance genes. Received: 7 July 1997 / Accepted: 6 October 1997     

A High Through-put Procedure for Capturing Microsatellites from Complex Plant Genomes

A method is outlined for large-scale isolation and characterization of microsatellite sequences from complex plant genomes. The method presented here differs from the previously published procedures in the use of randomly sheared (nebulized) genomic DNA for adapter-ligation, rigorous removal of biotinylated oligos, and high-density colony blots for constructing enriched libraries. Using this method we have constructed cotton microsatellite enriched libraries with over 20% (high stringency screening) or 75% (by random sequencing). Thus far we have identified and sequenced over 500 cotton microsatellites using this procedure. The procedure can be used to generate enriched SSR libraries from genomic DNA in about one week. High throughput screening and automated DNA sequencing can be accomplished in less than one month.     

Maintenance of the induced hypomethylated state of tobacco nuclear repetitive DNA sequences in the course of protoplast and plant regeneration

Previously, we established experimental conditions allowing us to induce hypomethylation of tandem arrays of highly repetitive DNA sequences in the Nicotiana tabacum L. nuclear genome (M. Bezdk et al. 1991, Planta 184, 487–490). In this paper, we demonstrate that loci containing the highly repetitive sequences of the HRS60 family can maintain the induced hypomethylated state through protoplast regeneration, non-differentiated callus growth, and plant regeneration. The hypomethylation, induced with 5-azacytidine and monitored on these sequences, did not substantially alter the capacity of calli to produce plants. It can be concluded that, in contradistinction of multiple copies of transgenes or ectopic genes which are usually recognized as methylation targets, endogenous tandem repeats, such as the HRS60, present at 10⁵ copies in the genome, can escape de-novo methylation.Abbreviation AzaC 5-azacytidine This project was supported by the Grant Agency of the Academy of Sciences of the Czech Republic. We thank Ms. Libue Jedliková, Ms. Hana Suchánková, and Ms. Emilie Koudelková for technical assistance.     

Chimpanzee microsatellite PCR primers applied to paternity testing in a captive colony

Previously designed primers for the polymerase chain reaction (PCR) amplifying microsatellite DNA segments containing GT/AC dinucleotide repeats in the chimpanzee (Pan troglodytes) genome were used for paternity testing in a breeding colony in captivity. Combinations of three PCR primers identified the fathers of all the tested 40 chimpanzees born in an eight-year period. The results suggested: (1) a positive (though not conclusive) correlation between male rank and number of offspring; (2) choice of mating partners by the female rather than by the male; and (3) absence of stable mating pairs over the years. For studies of chimpanzees in captivity and in the wild, these primers should be useful for paternity testing, for investigating genetic variations, and for improving genetic maintenance of breeding colonies.     

Structure and expression of an inhibitor of fungal polygalacturonases from tomato

A polygalacturonase inhibitor protein (PGIP) was characterized from tomato fruit. Differential glycosylation of a single polypeptide accounted for heterogeneity in concanavalin A binding and in molecular mass. Tomato PGIP had a native molecular mass of 35 to 41 kDa, a native isoelectric point of 9.0, and a chemically deglycosylated molecular mass of 34 kDa, suggesting shared structural similarities with pear fruit PGIP. When purified PGIPs from pear and tomato were compared, tomato PGIP was approximately twenty-fold less effective an inhibitor of polygalacturonase activity isolated from cultures of Botrytis cinerea. Based on partial amino acid sequence, polymerase chain reaction products and genomic clones were isolated and used to demonstrate the presence of PGIP mRNA in both immature and ripening fruit as well as cell suspension cultures. Nucleotide sequence analysis indicates that the gene, uninterrupted by introns, encodes a predicted 36.5 kDa polypeptide containing amino acid sequences determined from the purified protein and sharing 68% and 50% amino acid sequence identity with pear and bean PGIPs, respectively. Analysis of the PGIP sequences also revealed that they belong to a class of proteins which contain leucine-rich tandem repeats. Because these sequence domains have been associated with protein-protein interactions, it is possible that they contribute to the interaction between PGIP and fungal polygalacturonases.