Allele Frequency of 15 Short Tandem Repeats (Strs) in a Buganda Population (Central Uganda): Forensic Utility and Parentage Testing

Autosomal STR markers were first described as effective tools for human identity testing in the early 1990s [1] and have become the common currency of data exchange for human identity testing both in forensic casework and paternity testing [2]. Population databases allow for estimations of how rare or common a DNA profile may be in a particular population [3]. One population for which Short Tandem Repeats (STR) allele frequencies are not available is the Baganda tribe of Uganda. The aim of this work was to generate a Buganda population based 15 STR allele frequency data. We genotyped by multiplex PCR system using AmpF/STR® IdentifilerTM Plus PCR Amplification Kit that co-amplifies the 13 Combined DNA Index System (CODIS) STR loci plus additional two tetrameric markers; D2S1338 and D19S433, as well as the amelogenin locus for gender identification. In this study we present the allele frequencies and statistical data of forensic importance in the Buganda population.


Introduction
Autosomal STR markers were first described as effective tools for human identity testing in the early 1990s [1] and have become the common currency of data exchange for human identity testing both in forensic casework and paternity testing [2]. Population databases allow for estimations of how rare or common a DNA profile may be in a particular population [3]. One population for which Short Tandem Repeats (STR) allele frequencies are not available is the Baganda tribe of Uganda. The aim of this work was to generate a Buganda population based 15 STR allele frequency data. We genotyped by multiplex PCR system using AmpF/STR ® Identifiler TM Plus PCR Amplification Kit that co-amplifies the 13 Combined DNA Index System (CODIS) STR loci plus additional two tetrameric markers; D2S1338 and D19S433, as well as the amelogenin locus for gender identification. In this study we present the allele frequencies and statistical data of forensic importance in the Buganda population.

Materials and Methods
Ethical clearance was sought from the School of Biomedical Sciences Institutional Review Committee (IRC) College of Health Sciences of Makerere University, (Research file reference: SBS105) and Uganda National Council of Science and Technology (Research file reference: HS 1422). Blood samples were obtained from 221unrelated individuals in the 10 randomly selected districts of the Buganda-Central Uganda who had previously consented. DNA was extracted following Chelex Extraction protocol [4] and amplified using the AmpFlSTR ® Identifiler Plus TM PCR amplification kit following the manufacturer's user manual. Separation of the amplicons was done by capillary electrophoresis on an ABI 3130xl Genetic Analyzer using HiDi Formamide to fragment the bases and Gene Scan 500 Liz to size the fragmented bases following Applied Biosystems Instruction Manual (2006): Starting Electrophoresis. The resulting STRs were genotyped using the GeneMapper ® ID software V3.2. Allele frequency determination, Exact test probability for Hardy-Weinberg equilibrium (p-value), observed heterozygosity (Ho), expected heterozygosity (He) and inbreeding coefficient expressed as Wright's fixation index (F is ) were computed using GenePop v 4.1.3 software. Forensic and paternity efficiency parameters were computed using Promega Powerstat v12 excel spreed sheet.

Results and Discussion
The allele frequency of the 15 STRs generated of the Buganda population (Table 1) was below 50% at all loci as shown by the predominant alleles at each loci reflecting the usefulness and validity of these loci in calculating paternity indices and discriminating individuals [5]. All samples were successfully amplified and genotyped as shown by electropherogram images of the positive control; a quality control measure in genotyping process and selected samples of the Buganda population at the most heterozygous, polymorphic, and discriminating loci namely D21S11, D2S1338, D19S433 and FGA loci ( Figure 1). All loci were in Hardy-Weinberg equilibrium (p-values >0.05) with the observed heterozygosity very close to the expected heterozygosity. High levels of observed heterozygosity ranging from 69.23% (D7S820) to 87.78% (D21S11) is an indication that the Buganda population has a high level of genetic variation and this could be successfully utilized in discriminating between individuals [6]. Both the heterozygosity and Fis values indicate that Buganda population is a heterozygous population with limited genetic inbreeding if any [7] and this is attributed to cultural practice of intermarriages among different clans and prohibition of marriages within the same clan.
All loci were highly polymorphic with polymorphic information content (PIC) values ranging from 0.67 at D13S317 to 0.89 at D2S1338 all above 0.5 indicating good informativeness of all the tested STR markers and useful for identification purposes [6]. The power of discrimination (PD) across all tested loci was above 80% (from 0.875 at D13S317 to 0.978 at D2S1338) an indication that all loci are highly discriminating, that is, an innocent person will be excluded as the donor of an evidence unknown sample [6]. The most informative markers were   the most discriminating ones in the Buganda population as it was in the population of Island of Cres, Croatia [7]. The calculated combined match probability (CMP) of 1 in 2.471×10 17 and the combined power of discrimination (CPD) which was greater than 0.999999998 in the Buganda population will have an important role in the analysis of mixed DNA profiles such as in sexual assault cases as well as matching a control sample to the unknown sample [6].
The typical paternity index values at all loci were greater than 1, indicative of relatedness [8,9]. High paternity exclusion (PE) values ranging from 0.416 (D7S820) to 0.750 (D21S11) were observed for the Buganda population which showed that the loci could powerfully exclude a falsely accused individual as a biological father to a child and the higher the PE value, the more non-fathers are excluded [9]. Combined, the 15 loci yielded a combined typical paternity index (CTPI) of 565,852.41 and a combined power of paternity exclusion (CPE) of 0.99981 (greater than 99%) indicating a high power of exclusion [6].

Conclusion
The Buganda population is genetically diverse, all loci were very informative, highly polymorphic with high power of discrimination and exclusion. When combined, these loci are powerful genetic markers for forensic identification and paternity testing in the Buganda population.