Concordance study between the AmpFlSTR MiniFiler PCR amplification kit and conventional STR typing kits

The AmpFlSTR MiniFiler 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 MiniFiler 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 MiniFiler 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.     

Swiss Caucasian population data for the STR loci D2S1338 and D19S433 using the AmpFISTR SGM plus PCR amplification kit

Allele and genotype frequencies for the ten STR loci D3S1358, VWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, TH01, FGA were determined in a Swiss Caucasian population sample (n=206) using the AmpFISTR SGM Plus Amplification kit. Electrophoresis was carried out on an ABI PRISM CE 310 Genetic Analyzer instrument. Previously, allele frequencies were published for the 13 STR loci D3S1358, VWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820, THO1, TPOX, CSF1PO and D16S539 for the same samples (n=206) amplified with the AmpFISTR Profiler Plus and Cofiler PCR Amplification kits. Since the results for the eight loci D3S1358, VWA, FGA, D8S1179, D21S11, D18S51, THO1, D16S539 shared between the AmpFISTR SGM Plus, Profiler Plus and Cofiler PCR Amplification kits already are published, only the allele frequencies for the two STR loci D2S1338 and D19S433 are reported in this paper. The two loci meet Hardy-Weinberg expectations. In addition, there is little evidence for association of alleles among the 15 loci (amplified with the Profiler, Cofiler, and SGM Plus amplification kits). The allelic frequency data can be used in forensic analyses to estimate the frequency of a multiple STR locus DNA profile in the Swiss population.     

Developmental validation of a single-tube amplification of the 13 CODIS STR loci, D2S1338, D19S433, and amelogenin: the AmpFlSTR Identifiler PCR Amplification Kit

Analysis of length polymorphism at short tandem repeat (STR) loci utilizing the polymerase chain reaction (PCR) process has proven to be an ideal assay for human identification purposes. The short length of STR loci coupled with the amplification of target sequence through PCR allows for a robust, sensitive, and specific assay for highly polymorphic markers. A multiplex containing fifteen STR loci plus the gender-determining locus Amelogenin was developed to provide a single amplification/detection of all CODIS (Combined DNA Index System) STR loci (CSF1PO, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, D21S11, FGA, TH01, TPOX, and vWA) as well as two internationally-accepted STRs (D2S1338 and D19S433). By incorporating five-dye fragment analysis technology and non-nucleotide linkers, previously optimized AmpFlSTR kit primer sequences have been maintained. This kit has been developed in accordance with the standards of the forensic community as defined by the DNA Advisory Board. Validation studies were performed to include developmental validation, and the results support the use of the AmpFlSTR Identifiler PCR Amplification Kit for human identity and parentage testing.     

Developmental validation of the AmpFℓSTR® Identifiler® Plus PCR Amplification Kit: an established multiplex assay with improved performance

Abstract: Analysis of length polymorphism at short tandem repeat (STR) loci utilizing multiplex polymerase chain reaction (PCR) remains the primary method for genotyping forensic samples. The AmpF?STR^® Identifiler^® Plus PCR Amplification Kit is an improved version of the AmpF?STR^® Identifiler^® PCR Amplification Kit and amplifies the core CODIS loci: D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, D21S11, CSF1PO, FGA, TH01, TPOX, and vWA. Additional loci amplified in the multiplex reaction are the sex‐determinant, amelogenin, and two internationally accepted loci, D2S1338 and D19S433. While the primer sequences and dye configurations were unchanged, the AmpF?STR^® Identifiler^® Plus PCR Amplification Kit features an enhanced buffer formulation and an optimized PCR cycling protocol that increases sensitivity, provides better tolerance to PCR inhibitors, and improves performance on mixture samples. The AmpF?STR^® Identifiler^® Plus PCR Amplification Kit has been validated according to the FBI/National Standards and Scientific Working Group on DNA Analysis Methods (SWGDAM) guidelines. The validation results support the use of the AmpF?STR^® Identifiler^® Plus PCR Amplification Kit for human identity and parentage testing.     

STR data for the AmpFlSTR SGM Plus from the eastern and western sections of Mediterranean region of Turkey

Allele frequencies of the 10 STR loci (D16S539, D2S1338, D3S1358, vWA, D18S51, D21S11, D8S1179, D19S433, FGA and TH01) included in the AmpFlSTR SGM Plus kit were determined in 107 unrelated individuals from the eastern section and 108 unrelated individuals from western section of Mediterranean region of Turkey. The expected performance of these loci for personal identification and paternity testing in these populations were estimated.     

Population data for the AmpFl STR Identifiler PCR Amplification Kit in China Han in Jilin Province, China

Allele frequencies for the fifteen short tandem repeats (STR) loci D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820, D19S433, THO1, D16S539, CSF1PO, TPOX, D2S1338 (AmpFl STR Identifiler PCR Amplification Kit, Applied Biosystems) were determined in a population sample of unrelated China Han in Jilin Province, China. Statistical analyses were performed using Powerstats version 1.2 and GEP-ISFGWG. The results showed that all the loci met Hardy-Weinberg expectation.     

Validation of the AMPFℓSTR® MiniFilerTM PCR Amplification Kit for Use in Forensic Casework*

Abstract: The AmpF?STR^® MiniFiler^TM PCR Amplification Kit is designed to genotype degraded and/or inhibited DNA samples when the AmpF?STR^® Identifiler^TM PCR Amplification Kit is incapable of generating a complete genetic profile. Validation experiments, following the SWGDAM guidelines, were designed to evaluate the performance of MiniFiler. Data obtained demonstrated that MiniFiler, when used in conjunction with Identifiler, provided an increased ability to obtain genetic profiles from challenged samples. The optimum template range was found to be between 0.2 and 0.6 ng, with 0.3 ng yielding the best results. Full concordance was achieved between the MiniFiler kit and Identifiler kit except in a single case of a null allele at locus D21S11. Numerous instances of severe heterozygous peak imbalance (<50%) were observed in single source samples amplified within the optimum range of input DNA suggesting that caution be taken when attempting to deduce component genotypes in a mixture.     

Population data on the AmpF/STR Identifiler loci in Africans and Europeans from South Africa

Allele frequencies of 15 short tandem repeat (STR) loci, D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, vWA, TPOX, D18S51, D5S818 and FGA, were determined for 98 unrelated Africans from South Africa and 98 unrelated Europeans from South Africa using the AmpFlSTR Identifiler PCR amplification kit. The genotype frequency distributions of the 15 STR loci were in the Hardy-Weinberg equilibrium for both populations.     

STR data for the AmpFlSTR SGM Plus from Aegean region of Turkey

Allele frequencies of the 10 STRs loci (D16S539, D2S1338, D3S1358, vWA, D18S51, D21S11, D8S1179, D19S433, FGA, THO1) included in the AmpFlSTR SGM Plus Kit (PE Applied Biosystems) were obtained from a sample of 166 unrelated individuals in Aegean region of Turkey.     

Typeability of AmpFlSTR SGM Plus loci in brain and thyroid gland tissue samples incubated in different environments

Autolysis and putrefaction are crucial factors responsible for degradation of cells, tissues, and organs. Postmortem changes may assume different course depending on extrinsic and intrinsic conditions. The aim of the study was assessment of environmental effect on typeability of AmpFlSTR SGM Plus loci: D3S1358, VWA, D16S539, D2S1338, D81179, D21S11, D18S51, D19S433, TH01, FGA, and gender marker amelogenin. Brain and thyroid gland tissue specimens collected during autopsies of five persons aged 20-30 years were incubated at 21 degrees C and 4 degrees C in different environmental conditions. DNA was extracted by organic method from tissue samples collected in 7-day intervals and subsequently typed using AmpFlSTR SGM Plus kit and ABI 310. A fast decrease in typeability rate was seen in specimens incubated in peat soil and in sand. Brain tissue samples were typeable in all AmpFlSTR SGM Plus loci within 126 days of incubation at 4 degrees C. Faster DNA degradation was recorded in thyroid gland specimens. In samples with negative genotyping results, no DNA was found by fluorometric quantitiation.     

STR data for the AmpFlSTR SGM Plus from Marmara region of Turkey

Allele frequencies of the 10 STRs loci (D16S539, D2S1338, D3S1358, vWA, D18S51, D21S11, D8S1179, D19S433, FGA, THO1) included in the AmpFlSTR SGM Plus kit (PE Applied Biosystems) were obtained from a sample of 173 unrelated individuals in Marmara region of Turkey.     

A single mutation in the FGA locus responsible for false homozygosities and discrepancies between commercial kits in an unusual paternity test case

We report an unusual paternity test case showing multiple peculiarities. Using AmpFlSTR Profiler Plus and AmpFlSTR Identifiler PCR Amplification kits, the alleged father and the two children were apparently homozygous at the FGA locus, but using the PowerPlex 16 kit the three individuals were found to be heterozygous. Drop-out was caused by a single mutation event in the presumptive binding site of the reverse primer. In addition, three inconsistencies were detected between the daughter and the alleged father among 18 STR markers. The occurrence of the rare null allele at the FGA locus and case history suggested that the true father was the brother of the alleged father. Furthermore, a single-step repeat maternal mutation was also detected at D16S539. This puzzling case was solved by using multiple analytical approaches, including the use of different primer pairs, the use of a high number of STR markers, and the characterization of the mutation causing the "null allele."     

Autosomal short tandem repeat analysis of ancient DNA by coupled use of mini- and conventional STR kits

Abstract: Multiplex autosomal short tandem repeat (STR) genotyping enables researchers to obtain genetic information from ancient human samples. In this study, we tested newly developed AmpF?STR^® MiniFiler? kit for autosomal STR analysis of ancient DNA (aDNA), using human femurs (n = 8) collected from medieval Korean tombs. After extracting aDNA from the bones, autosomal STR analyses were repeated for each sample using the AmpF?STR^® MiniFiler? and Identifiler? kits. Whereas only 21.87% of larger‐sized loci profiles could be obtained with the Identifiler? kit, 75% of the same loci profiles were determined by MiniFiler? kit analysis. This very successful amplification of large‐sized STR markers from highly degraded aDNA suggests that the MiniFiler? kit could be a useful complement to conventional STR kit analysis of ancient samples.     

Population data on the AmpFlSTR SGM plus PCR amplification kit in Germans and Austrians

Allele frequencies for the short tandem repeat loci D3S1358, VWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, THO1 and FGA were determined in 231 German and 100 Austrian unrelated Caucasoids using the AmpFlSTR SGM plus kit (Applied Biosystems). All loci met Hardy-Weinberg expectations. In 212 meioses per locus, one mutation event for D19S433 was observed.     

Allele distribution of 15 PCR-based loci in the Rwanda Tutsi population by multiplex amplification and capillary electrophoresis

A genetic study of 15 autosomal STRs is carried out (D2S1338, D3S1358, D5S818, D7S820, D8S1 79, D13S317, D16S359, D18S51, D19S433, D21S11, CSF1PO, FGA, TPOX, THO1, VWA) in a sample of unrelated Tutsis. The molecular phenotypes were determined by means of multiplex strategies (AmpFlSTR Identifiler PCR Amplification Kit, Applied Biosystems) followed by capillary electrophoresis.     

STR data for the AmpFlSTR SGM Plus and Profiler loci from Taiwan

Allele frequencies for the 15 STRs included in the AmpFlSTR SGM Plus and Profiler kit (D3S1358, VWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, THO1, FGA, TPOX, CSF1PO, D5S818, D13S317 and D7S820) were obtained from a sample of 597 unrelated individuals living in Taiwan. All loci, except for D8S1179 and D5S818, are in Hardy-Weinberg equilibrium.     

Population genetics of 10 STR loci in a population of Central Poland

Allele frequencies for the 10 short tandem repeats (STRs) included in AmpFlSTR SGM Plus amplification kit (D3S1358, vWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, TH01, FGA) were estimated from a sample of 500 unrelated individuals born in Central Poland.     

3种常用PCR扩增试剂盒检验血样DNA的结果比较

目的探讨常用的ProfilerPlusTM、IdentifilerTM与Powerplex(16试剂盒检验血样DNA的差异。方法510名无关中国汉族个体血样,分别用ProfilerPlusTM与Powerplex(16试剂盒进行DNA检验,然后对有不同检验结果的同一样本,再用ProfilerPlusTM、IdentifilerTM与Powerplex(16等三种试剂盒进行检验,并比较其结果。结果在510名个体血样的DNA检测结果中,发现同一样本有不同结果的有7例,其差异率为1.3725%;ProfilerPlusTM、IdentifilerTM与Powerplex(16各有1例在D13S317或FGA基因座上出现等位基因缺失现象,缺失率为0.1961%;ProfilerPlusTM有5例在D8S1179基因座上出现扩增严重不平衡现象,相应的IdentifilerTM与Powerplex(16试剂盒的检验结果为正常杂合子。结论ProfilerPlusTM、IdentifilerTM与Powerplex(16试剂盒检验血样DNA均会出现扩增不平衡和/或基因丢失现象,其发生几率IdentifilerTM与Powerplex(16试剂盒较ProfilerPlusTM少。     

荧光标记3个miniSTR基因座复合扩增系统的建立

目的建立扩增片段<135bp,包括D5S818,D8S1179,D16S539 3个miniSTR基因座复合扩增系统。方法采用不同荧光染料标记引物,通过PCR扩增,利用ABI 3100遗传分析仪进行片段长度分析,对100份无关个体血样,10个家系样本以及30份高度降解检材进行检测。结果本系统DNA分型结果与AmpFLSTR Identifiler试剂盒完全一致,且灵敏度高于AmpFLSTR Identifiler试剂盒。结论本系统可以应用于个人识别和亲权鉴定,为降解DNA样本分型提供了新的方法。     

Maine Caucasian population DNA database using twelve short tandem repeat loci

A population study of Caucasians residing in Maine was conducted using the AmpF1STR Profiler PCR Amplification Kit and the AmpF1STR Profiler Plus PCR Amplification Kit (Applied Biosystems Division (ABD) of Perkin Elmer, Foster City, CA). The kits contain the reagents necessary to amplify 12 different STR loci and the gender marker Amelogenin using two multiplex PCR, each containing nine STR loci. Thus, there is an overlap of six STR loci. The 12 STR loci are TH01, TPOX, CSF1PO, D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, and D7S820. These loci represent 12 of the 13 core loci selected by the CODIS STR standardization project. Dye-labeled amplification products were separated and detected using the capillary electrophoresis instrument ABI Prism 310 Genetic Analyzer. Allele frequencies were determined for the 12 STR loci. Statistical analysis of the data included Hardy-Weinburg equilibrium (HWE) analysis, pairwise independence testing, power of discrimination (PD), and probability of exclusion (PE).