Concordance study and population frequencies for 16 autosomal STRs analyzed with PowerPlex® ESI 17 and AmpFℓSTR® NGM SElect™ in Somalis, Danes and Greenlanders.

A concordance study of the results of PowerPlex(®) ESI 17 and AmpFℓSTR(®) NGM SElect™ kits obtained from 591 individuals from Somalia (N=198), Denmark (N=199) and Greenland (N=194) was performed. Among 9456 STR types, seven discordant results were found with the two kits: one observed in the D19S433 system in an individual from Denmark and six in the SE33 system in six individuals from Somalia. Sequencing of SE33 in the six samples with discordant results showed G>A transition 15bp downstream of the repeat unit in three of the individuals, and G>A transition 68bp downstream of the repeat unit in the other three individuals. Population data for 16 autosomal STR systems analyzed in 989 individuals from Somalia, Denmark and Greenland are also presented. The highest mean heterozygosity was observed in Danes (82.5%). With the exception of D8S1179 in Danes, no significant deviations from Hardy-Weinberg expectations were observed. Only one pair of systems (D12S391 and D18S51) showed significant allelic association in Greenlanders (after Holm-Sidák correction). A MDS plot drawn from pairwise FST values calculated between 21 populations showed a clear displacement of the Greenlandic population versus the other ones included in the analyses. The highest combined chance of exclusion and power of discrimination was observed for Danes reaching values of 99.9999987% and 1 in 1.8×10(21), respectively.

Effect of mutations in the upstream promoter on the transcription of human 5S rRNA genes.

The human 5S rRNA gene has a 12-mer external promoter, the D box, localized about 30 bp upstream the coding sequence. By site directed mutagenesis 58 different D box promoter mutants were made. While some mutations in the D box allowed full transcription, other mutations decreased the transcriptional activity to 20-50% compared to the bona fide gene, showing the importance of this external promoter in transcription initiation. A number of maxi 5S rRNA genes were constructed from bona fide genes and D box mutated clones. Transfection of HeLa cells with maxi 5S rRNA genes showed that the D box is also important for 5S rRNA gene expression in vivo. Evidence from different eukaryotic cells suggests that expression of 5S rRNA genes is regulated by external promoters in addition to the internal control region.

A report of the 1997, 1998 and 1999 Paternity Testing Workshops of the English Speaking Working Group of the International Society for Forensic Genetics.

We present the results of the 1997, 1998 and 1999 Paternity Testing Workshops of the English Speaking Working Group of the International Society for Forensic Genetics. The numbers of participating laboratories were 24 (1997), 31 (1998) and 32 (1999). In 1997, all laboratories drew the correct conclusion that the alleged father was the biological father of the child. In 1998, the alleged father was the biological brother of the child and all laboratories excluded him. The scenario in 1999 was a deficiency case consisting of mother, child and the parents of the alleged father and all but one laboratory drew the correct conclusion.The percentage of laboratories routinely performing variable number of tandem repeat (VNTR) investigations using single locus probes (SLPs) and restriction fragment length polymorphism (RFLP) decreased from 83% in 1997 to 66% in 1999. In the three workshops, more than 90% of the laboratories used polymerase chain reaction (PCR) based systems. In 1999, 80% of the laboratories performing PCR, used commercially available short tandem repeat (STR) kits. Other commonly used systems were HLA and PolyMarker investigated with PCR. Conventional systems and RFLP investigations of VNTRs with multi loci probes (MLPs) were used routinely by approximately 20% of the participating laboratories. All laboratories submitting results in the three workshops used RFLP-based VNTRs or/and PCR based VNTRs/STRs. Inter-laboratory comparisons of the results showed a very high concordance. The overall coefficients of variation between the laboratories of the results of RFLP typing of the commonly used VNTRs D2S44, D7S21, D7S22 and D12S11 were 1.2-1.3%. Consistent results were obtained in the great majority of the systems investigated by PCR and typing errors counted for less than 0.3% of the PCR based results.

Mink 5S rRNA genes map to 2q in three loci suggesting conservation of synteny with human 1q.

By in situ hybridization we show that the SS rRNA genes in the mink map to chromosome 2q in three loci. The 2q1.1 locus containing 34% of the 5S rDNA, maps close to the centromere, and the remaining two loci of the 5S rDNA map to 2q1.3 (52%) and to 2q2.3. (14%). These data were obtained with a tritiated transcript of the 5S rRNA gene containing 121 bp. In a comparative FISH study performed with a biotinylated transcript of the 5S rRNA gene the procedure failed to detect the 2q2.3 site. A closely corresponding difference between the two procedures experienced previously in man and in the crab-eating macaque is discussed. The present results suggest a homology between 2q in the mink and part of 1q in man harbouring the 5S rRNA genes in 1q42.13 and 1q31, respectively.

The rat 5S rRNA bona fide gene repeat maps to chromosome 19q12-->qter and the pseudogene repeat maps to 12q12.

The bona fide 5S rRNA genes in the rat are found in a 1.8-kb tandem repeat and the pseudogenes occur in a 2.5-kb tandem repeat. Three bona fide 5S rRNA genes and one gene variant with one base substitution in the coding region were isolated from the 1.8-kb repeat. Six pseudogenes were isolated from the 2.5-kb repeat. The total number of genes/gene variants/pseudogenes is 700-1200 copies per haploid genome, and the pseudogene repeat contains about 50% more 5S rDNA related sequences compared with the bona fide gene repeat. Various well-defined 5' - and 3'-flanking sequences of the bona fide gene and of the pseudogene were used for in situ hybridization to metaphase chromosomes. The results showed that the bona fide 5S rRNA gene repeat Rn5s maps to chromosome 19q12 and the pseudogene repeat Rn5sp maps to 12q12.

Additional assignment of the human 5S rRNA genes to chromosome region 1q31.

A major fraction of the human 5S rRNA genes has previously been assigned to chromosome region 1q42.11-->q42.13 (Sørensen et al., 1991). Through in situ hybridization of different tritiated probes to metaphase spreads, we have demonstrated that a minor fraction of the 5S rRNA genes is localized at band 1q31. This fraction amounts to 25-30% of the genes found in the region 1q42.11-->q42.13. Results obtained with the chromosomes of a balanced translocation carrier involving this region indicate that the major cluster is localized in band 1q42.13.

Characterization of 5S rRNA genes from mouse.

In order to characterize the transcriptional regulation of the 5S rRNA genes we have isolated a bona fide gene and a pseudogene from mouse cells. These 5S rRNA genes contain a 12-bp sequence designated as the D-box, located in position -33 to -22 bp, and two Sp1-binding sites in the 5'-flanking region. The D-box is conserved in human and hamster 5S rRNA genes although in slightly different upstream positions. The bona fide mouse 5S rRNA gene was transcribed in a HeLa S-100 extract. The transcriptional activity of this gene was only 50% of that of the human gene, indicating the involvement of species-specific transcription factors and/or polymerases. The pseudogene which contains the D-box, but with position +25 to +35 bp deleted, showed no transcriptional activity. Deletion of the D-box in the 5'-flanking sequence abolished transcriptional activity, indicating that this conserved sequence is of importance for gene expression.

Transcription of human 5S rRNA genes is influenced by an upstream DNA sequence.

Six human 5S rRNA genes and gene variants and one pseudogene have been sequenced. The six genes/variants were transcribed in a HeLa cell extract with about equal efficiency. Three genes contain the Sp1 binding sequence GGGCGG in position -43 to -38 and three genes contain the Sp1 like sequence GGGCCG in this position. The six genes contain furthermore one Sp1 binding site in a position about -245 and one ATF recognition site in a position about -202. A 12 bp sequence (GGCTCTTGGGGC) found in position -32 to -21 strongly influenced the transcriptional efficiency in vitro. This 12-mer, designated the D box, has also been found upstream a 5S rRNA gene from hamster and mouse. Removal of the Sp1 binding sites had no effect on the transcription in vitro whereas the transcriptional efficiency decreased to 10% if the D box was removed from the human 5S rRNA gene.