Article

Concordance study between the AmpFlSTR((R)) MiniFiler (TM) PCR amplification kit and conventional STR typing kits

Journal of Forensic Sciences (Impact Factor: 1.16). 06/2007; 52(4):870 - 873. DOI: 10.1111/j.1556-4029.2007.00491.x

ABSTRACT

The AmpFℓSTR® MiniFilerTM polymerase chain reaction amplification kit developed by Applied Biosystems enables size reduction on eight of the larger STR loci amplified in the Identifiler® kit, which will aid recovery of information from highly degraded DNA samples. The MiniFilerTM Kit amplifies CSF1PO, FGA, D2S1338, D7S820, D13S317, D16S539, D18S51, and D21S11 as well as the sex-typing locus amelogenin. A total of 1308 samples were evaluated with both the MiniFilerTM and Identifiler® STR kits: 449 African American, 445 Caucasian, 207 Hispanic, and 207 Asian individuals. Full concordance between Identifiler and MiniFiler Kits was observed in 99.7% (10,437 out of 10,464) STR allele calls compared. The 27 differences seen are listed in Table 1 and encompass the loci D13S317 (n = 14) and D16S539 (n = 10) as well as D18S51 (n = 1), D7S820 (n = 1), and CSF1PO (n = 1). Genotyping discrepancies between the Identifiler and MiniFiler kits were confirmed by reamplification of the samples and further testing using the PowerPlex® 16 kit in many cases. DNA sequence analysis was also performed in order to understand the nature of the genetic variations causing the allele dropout or apparent repeat unit shift.
Sample sources include IBB, MLN, DDC, and ABI. With only three exceptions (see samples no. 1, 2, 27), PowerPlex® 16 (PP16) results agree with the Identifiler® results for these samples. DNA sequencing was performed to ascertain the genetic variation responsible for the discordance of the impacted allele (shown in bold font). Note that sample no. 15 is the child of sample no. 14, sample no. 20 is the child of sample no. 21, and sample no. 22 is the child of sample no. 23.
AA, African American; C, Caucasian; H, Hispanic; A, Asian; MLN, Millennium; IBB, Interstate Blood Bank; DDC, DNA Diagnostic Center; ABI, Applied Biosystems.

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Available from: Julio J Mulero, Apr 07, 2014
    • "In addition, a 4 bp deletion (rs561167308, 0.4 % frequency in 1000 Genomes Phase 3 dataset) has been reported 24 bases downstream from the core TATC repeat [84] [85]. This deletion can impact lengthbased allele designations with different primer sets, as seen by Hill et al. [24]. Similar impacts may be observed with sequencing applications. "
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    ABSTRACT: The development of next generation sequencing (NGS) technologies creates the potential for changing the method by which the forensic science community genotypes short tandem repeat (STR) loci. While the capabilities of NGS are promising, moving from current capillary electrophoresis (CE) methods would require new guidelines to be established and a new understanding of artifacts that may arise with the use of NGS. Stutter has been well characterized for CE technologies; however, NGS workflows may use different polymerases and amplification approaches, which could alter the appearance of this artifact. Stutter is most commonly seen in the n-4 position in CE data, but may be observed more rarely in the n+4 and n-8 positions. NGS data frequently contains detectable sequences consistent with stutter at the n+4 and n-8 positions, and may even contain stutter at the n-12 position for some loci. It is possible that these alternate types of stutter events occur at similar levels in CE workflows and go undetected due to the analytical threshold employed or because the artifacts do not exceed the background noise. Comparing stutter events in NGS data to what has been observed by CE will improve our understanding of the effects of library preparation and sequencing. Characterizing stutter events by sequence will contribute to the development of guidelines and facilitate implementation of NGS technology. Further, determining stutter ratios for each isoallele would allow for individual sequence thresholds to be set, which could then be used to improve mixture interpretation models.
    No preview · Article · Sep 2015 · Forensic Science International Genetics Supplement Series
    • "In addition, a 4 bp deletion (rs561167308, 0.4 % frequency in 1000 Genomes Phase 3 dataset) has been reported 24 bases downstream from the core TATC repeat [84] [85]. This deletion can impact lengthbased allele designations with different primer sets, as seen by Hill et al. [24]. Similar impacts may be observed with sequencing applications. "
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    ABSTRACT: This article reviews what is currently known about short tandem repeat (STR) allelic sequence variation in and around the twenty-four loci most commonly used throughout the world to perform forensic DNA investigations. These STR loci include D1S1656, TPOX, D2S441, D2S1338, D3S1358, FGA, CSF1PO, D5S818, SE33, D6S1043, D7S820, D8S1179, D10S1248, TH01, vWA, D12S391, D13S317, Penta E, D16S539, D18S51, D19S433, D21S11, Penta D, and D22S1045. All known reported variant alleles are compiled along with genomic information available from GenBank, dbSNP, and the 1000 Genomes Project. Supplementary files are included which provide annotated reference sequences for each STR locus, characterize genomic variation around the STR repeat region, and compare alleles present in currently available STR kit allelic ladders. Looking to the future, STR allele nomenclature options are discussed as they relate to next generation sequencing efforts underway. Published by Elsevier Ireland Ltd.
    No preview · Article · Jul 2015 · Forensic Science International: Genetics
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    • "The same applies to primer binding site mutations, which can result in a loss of amplification of a parentally inherited allele. These mutations are usually older and can be resolved after amplification with alternative primers [16] [17] [18]. "
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    ABSTRACT: Well defined estimates of mutation rates are a prerequisite for the use of short tandem repeat (STR-) loci in relationship testing. We investigated 65 isolated genetic inconsistencies, which were observed within 50,796 allelic transfers at 23 STR-loci (ACTBP2 (SE33), CD4, CSF1PO, F13A1, F13B, FES, FGA, vWA, TH01, TPOX, D2S1338, D3S1358, D5S818, D7S820, D8S1132, D8S1179, D12S391, D13S317, D16S539, D17S976, D18S51, D19S433, D21S11) in Caucasoid families residing in Austria and Switzerland. Sequencing data of repeat and flanking regions and the median of all theoretically possible mutational steps showed valuable information to characterise the mutational events with regard to parental origin, change of repeat number (mutational step size) and direction of mutation (losses and gains of repeats). Apart from predominant single-step mutations including one case with a double genetic inconsistency, two double-step and two apparent four-step mutations could be identified. More losses than gains of repeats and more mutations originating from the paternal than the maternal lineage were observed (31 losses, 22 gains, 12 losses or gains and 47 paternal, 11 maternal mutations and 7 unclear of parental origin). The mutation in the paternal germline was 3.3 times higher than in the maternal germline. The results of our study show, that apart from the vast majority of single-step mutations rare multi-step mutations can be observed. Therefore, the interpretation of mutational events should not rigidly be restricted to the shortest possible mutational step, because rare but true multi-step mutations can easily be overlooked, if haplotype analysis is not possible.
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