Genetic Testing for Additional Evidence during Investigations: Focus in Ethics / 대한법의학회지

Since the introduction of genetic fingerprinting 30 years ago, there has been considerable development in the field of forensic genetics. A cautious approach is emphasized when using human genetic evidence in order to protect individual rights and because of its distinctiveness. Nevertheless, conventional ethical guidelines may no longer be suitable for handling information derived from genetic material. Moreover, projected innovations to maximize such systems' usage have raised previously debated ethical concerns. Recent and on-going research on the use of genetic evidence obtained from crime scenes to estimate physical appearance, ancestry, and/or personal traits is expected to provide additional investigative resources, especially for cases involving unknown identity. Given the special nature of genetic components, ethical issues need to be seriously considered and addressed when conducting research involving human genetic material. However, such ethical parameters may shift with scientific advancements. Moreover, because ethics reflects social consensus, various perspectives must be obtained and discussed. This paper introduces multiple perspectives on using genetic material as additional evidence for police investigations and indicates scope for the discussion of prospective ethical concerns.

Analysis of Kinship Index Distributions in Koreans Using Simulated Autosomal STR Profiles / 대한법의학회지

Kinship testing in forensic casework is largely based on a likelihood ratio (LR) approach with short tandem repeat (STR) markers; however, in order to efficiently identify potential relatives in a specific population, the threshold values for kinship prediction should be determined by analyzing the kinship index distributions of the population in question. In this study, 250,000 DNA profiles were simulated using allele frequencies at 20 autosomal STR loci in Koreans, then the LRs were calculated for true close relatives and unrelated pairs. The LR distributions in related and unrelated pairs under a given relationship were compared in 2 sets of 13 Combined DNA Index System (CODIS) and 20 STR profiles. Using 13 CODIS STRs, true relatives in parent/child and full-sibling relationships were sufficiently discriminated from unrelated pairs with LR thresholds of 1,000 and 100, respectively. However, the CODIS STRs lacked the discriminatory power to differentiate between related and unrelated pairs in uncle/nephew and first cousin relationships due to high false-positive and false-negative rates with a LR threshold of 10. Increasing the number of STR loci to 20 increased discrimination of close relatives, but high false results remained in uncle/nephew and first cousin relationships. The kinship index data from this study will help make decisions on various kinship testing and familial searching in Koreans.

DNA Methylation-Based Age Estimation in the Forensic Field / 대한법의학회지

The estimation of age is an important issue in forensic science, and the forensic community has attempted many times to establish methods for solving this issue. Aging leads to alterations in tissues and organs at the molecular level. These alterations at the molecular level may aid forensic scientists to estimate the age of a living person or a dead body. Initially, the focus was on the genetic components of aging, but recently, epigenetic mechanisms have emerged as the key contributors to the alterations in genome structure and function that accompany aging. In particular, DNA methylation is one of the best-understood mechanisms, and it has been suggested as a promising biomarker for age estimation in many studies. In this review, we summarize the recent studies on age-associated DNA methylation changes in different tissues and discuss its possible and practical applications in forensics.

Rapid and Simple Screening of Mitochondrial DNA in Koreans by the Analysis of Highly Variable Control Region SNPs / 대한법의학회지

Human mitochondrial DNA (mtDNA) is generally used to identify highly degraded forensic samples, particularly when the extracted DNA is not sufficient for nuclear DNA analysis. However, direct sequencing, the most widely used mtDNA analysis method, is laborious and time-consuming, and precludes the simultaneous analysis of many samples. Here, we describe a rapid and simple screening method for mtDNA analysis in Koreans using single base extension (SBE) methods. Sixteen highly polymorphic mtDNA SNPs from the control region were selected, and a multiplex SBE system was constructed to analyze them. Because the developed system consists of two duplex PCRs, which produce small amplicons with fewer than 270 bp, it works well with highly degraded samples such as old skeletal remains. Using this multiplex SBE system, 145 different haplotypes were expected to be observed from 593 unrelated Koreans. Seventy-three haplotypes were expected to be observed only once, and the most frequent haplotype was expected to occur 80 times. Since the mean number of pairwise differences was estimated to be 4.55, the developed system could be useful to exclude samples that do not match evidence and reference samples. Therefore, the multiplex SBE system used in this study will be a useful tool to analyze many samples simultaneously and to efficiently screen out non-matching mtDNA sequences in forensic casework.

Web-based Y-STR Database for Haplotype Frequency Estimation and Kinship Index Calculation / 대한법의학회지

We developed a web-based Y chromosomal short tandem repeat (Y-STR) database (ySTRmanager, http://ystrmanager.yonsei.ac.kr) to facilitate calculation of Y-STR haplotype frequency estimates for random matches and kinship indices for various relationship levels. The ySTRmanager database provides 3 functions: (i) Y-STR haplotype search, (ii) kinship index calculation, and (iii) user database configuration. The Y-STR haplotype search function allows researchers to retrieve Y-STR haplotypes that meet queried Y-STR allele, Y-haplogroup affiliation, and/or sample information from a selected population in the open database, which consists of 12-17 Y-STR loci. The number of matches in a selected population, haplotype frequency estimator, and detailed results for matched and neighbor haplotypes are displayed as a set of search results. The kinship index calculation function provides kinship indices of 2 input Y-STR haplotypes for the relationship represented by the number of meioses, with consideration of target population and mutation rate of each Y-STR. In addition, ySTRmanager allows registered users to configure their own database to store and analyze Y-STR haplotype and/or mutation rate data. The stored Y-STR data can be used in the search function and in the analysis to obtain forensic statistical values. The ySTRmanager will be a useful system to analyze and manage Y-STR data in the practice of forensic genetics.

DNA Database Searching Using Genetic Relationship / 대한법의학회지

The usefulness of DNA databases is well known. In Korea, many cases have been solved since the establishment of DNA databases in 2010. DNA profiles obtained from criminal evidence are analyzed and are kept in databases, and matching profile is searched. On the matching occasion, DNA databases can provide some investigative information. Close family members, for example father, son or brother, share more alleles than unrelated people. This genetic closeness can be searched using the so called familial searching, and many successful cases have been reported. Management of DNA databases including familial searching needs continuous monitoring and control, especially from ethical view points. Many different views for familial searching have been presented, and each country running DNA databases has their own different policies. We present the utility, approaches and different policies of familial searching as well as arguments for and against familial searching. We also suggest that our appropriate guidelines be mandatory and a proper administrative process be prepared for the start of familial searching.

Application of a Repair Enzyme to STR Analysis of Damaged DNA / 대한법의학회지

DNA found at the scene of a crime is often damaged and degraded to small fragments and remained in low quantity. Therefore, there are some difficulties in forensic STR typing of these DNA samples. In this study, to overcome these limitations as different approach, we applied repairing enzyme to damaged DNA. The efficacy of a repair enzyme system (PreCR Repair Mix) was evaluated by using several types of artificially damaged and naturally damaged DNA. The results showed that autosomal STR amplification produced increased yield in the DNA damaged by UV irradiation, oxidation, or acid/heat, and DNA from dried blood spot and dried saliva spot by treatment of a repair enzyme, but not in DNA extracted from old skeletal remains. In conclusion, a repair enzyme will be efficiently applied to forensic samples which were damaged by UV irradiation, oxidation, and acid/heat.

RNA for Body Fluid Identification in Forensics / 대한법의학회지

Lately, there seems to be a growing interest in the forensic community for RNA analysis. Especially, body fluid identification using cell-specific RNA expression profiles is expected to present a new technique that will supplement DNA analysis in forensic casework. Several RNA markers specific to blood, saliva, semen, menstrual blood and vaginal secretions have been identified and their specificities and sensitivities have been confirmed using various forensic samples. Therefore, this review provides an overview of the present knowledge and the most recent developments in RNA analysis for the body fluid identification and discusses its possible and practical use in forensics.

Statistical Interpretation in Making DNA-based Identifications of Mass Victims / 대한법의학회지

DNA profiles have been increasingly used as the most reliable means to identify remains from war or mass disaster. To establish the identity with such a large set of victims, special care should be taken to correlate remains with correct family references while avoiding coincidental match between non-relatives. Therefore we address here relevant statistical and combinatorial issues in the DNA identification of mass victims. A simple and general formula for the likelihood ratio governing any potential kinship between two DNA profiles was presented, and for that purpose, the probabilities that a given relative and an individual share autosomal identical-bydescent alleles were calculated. In addition, a method dealing with the allele drop-out in kinship analysis and the estimation of a cold hit were discussed.

Genetic characteristics of 22 Y-STR loci in Koreans / 대한법의학회지

To construct a Korean Y-chromosomal STR database for 22 Y-STRs (DYS19, DYS385, DYS388, DYS389I/II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, DYS446, DYS447, DYS448, DYS449, DYS456, DYS458, DYS464, DYS635, and GATA H4.1), 708 DNA samples from unrelated Korean males were analyzed using three multiplex PCR systems. During analysis, thirty two microvariant alleles were observed at DYS449, DYS458, and DYS464, and duplicated alleles were occurred once each for DYS19, DYS390, and DYS447. In haplotype analysis for the 22 Y-STRs, a total of 693 different haplotypes were observed with overall haplotype diversity of 0.9999, and of these, 680 haplotypes were unique. By combining each marker for the extended SWGDAM haplotype, DYS458 was the most informative marker. In addition, the diversity of combined haplotypes of DYS447, DYS458, DYS635, GATA H4.1, and the SWGDAM Y-STR loci was comparable to haplotypes of 17 loci in the AmpFlSTR(R) Yfiler(TM) kit.

Evaluation of Nine Non-CODIS MiniSTR Loci to aid Analysis of Degraded DNA / 대한법의학회지

For highly degraded DNA samples of forensic casework, new miniSTR PCR systems have been developed to supplement the current CODIS STRs. In the present study, we established the three miniplexes for nine miniSTRs (NC01 : D10S1248, D14S1434 and D22S1045; NC02 : D1S1677, D2S441 and D4S2364; and NC03 : D3S3053, D6S474 and D20S482) which had been previously suggested by Butler group (NIST, Gaitherburg, MD, USA). To evaluate the usefulness of the nine miniSTRs in analysis of degraded DNA, the sensitivity and efficacy of the three miniplexes were determined and then compared with those of the BigMini STR system which consists of six CODIS miniSTRs (TH01, CSF1PO, FGA, TPOX, D7S820, and D21S11). The three miniplexes gave better results in both the sensitivity test and efficiency test in comparison with BigMini. In the sensitivity test using serially diluted standard DNA, most loci in the three miniplexes showed reliable results for samples containing 50 pg of DNA and some even showed good sensitivity for samples containing 30 pg of DNA. Additionally, the three miniplexes generated useful profiles for both enzymatically degraded DNA and 50-year old skeletal remain samples. Among the nine miniSTRs, D4S2364, D3S3053, D14S1434, and D1S1677 produced the most successful DNA profiles for old skeletal remains. These results suggest that new miniSTRs could be useful supplements to the 13 CODIS STRs for forensic analysis of degraded DNA.

Haplotypes and mutations of 17 Y-STR loci from Korean father-son pairs / 대한법의학회지

We have investigated 17 Y-STR loci (DYS19, DYS385a/b, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, DYS448, DYS456, DYS458, DYS635 (Y GATA C4), Y GATA H4) in 365 Korean father-son pairs of 355 families. Of 338 different haplotypes obtained from 355 fathers, 326 haplotypes were observed once, 10 haplotypes two times and the other two haplotypes were observed 4 and 5 times, respectively. The overall haplotype diversity was 0.9996. In 365 father-son pairs, a total of 21 mutations were observed at 12 Y-STR loci. Sequence analysis for mutant alleles demonstrated 21 single step mutations: 8 gains and 13 losses. However, there was no significant surplus of gains or losses. The locus-specific mutation rate estimates were between 0.0 and 8.2 x 10(-3) and the average mutation rate estimates were 3.4 x 10(-3)(95% C.I. 2.1-5.2 x 10(-3)) across all 17 Y-STR loci.

Intra-individual Difference of Length Heteroplasmy in Blood and Hair Shaft Mitochondrial DNA / 대한법의학회지

To observe mtDNA length heteroplasmy in a homoploymeric cytosine tract of the mitochondrial HV2 region, we carried out size-based separation of PCR products, which was produced by using primers designed to minimize the stutter production. Blood and hair shaft samples were collected from 25 individuals. The result showed significant qualitative/quantitative peak pattern variations among blood and hair shaft mtDNA profiles. Based on the results of this study, an exclusion depended solely on differences in length of the major C-tract variant could thus be an erroneous interpretation. Therefore, differences in the number of cytosine or qualitative/quantitative peak pattern variations in the C-tract of the mtDNA HV2 region cannot be used alone to support an interpretation of exclusion.

A Population Study of X-chromosomal STRs, DXS9898, DXS6809, DXS7424 and DXS10011 in Koreans / 대한법의학회지

A population study of the X-chromosomal short tandem repeat (STR) loci DXS9898, DXS6809, DXS7424 and DXS10011 was carried out by single multiplex PCR in a sample of 300 unrelated Korean individuals (150 males and 150 females). For accurate and reproducible STR typing, sequenced allelic ladders were constructed and GenoTyper macro was programmed. In this study, four types of the repre-sentative repeat sequence structure of DXS10011 were observed and the allele loss at DXS9898 was observed in 13 of 450 chromosomes (2.9%). The inter-population comparison of the allele frequencies at the 4 X-STRs showed significantly different distributions (p<0.01) for Koreans and Germans except DXS10011. All statistical parameters for forensic efficiency showed that the 4 X-chromosomal STRs are highly informative. Especially, DXS10011 is expected to be the most useful marker for forensic practice.

Simple Quantification and DNA Profiling from Degraded Low Copy Number DNA Samples / 대한법의학회지

DNA quantification is important to ensure the consistency and the reliability in the interpretation of degraded low copy number DNA typing. We applied the simple PCR quantification method using fluo-rescently labeled primers for the amplification of mtDNA and amelogenin gene in 50 year old skeletal remains (e.g. bone and tooth). K562 DNA was serially diluted and used as a standard for concentration marker to gauge the amount of DNA from PCR versus the peak area. The quantities of DNA extracted from bones and teeth did not show significant difference in the analyses both using mtDNA and amelo-genin gene as an amplification target. To test the efficiency of DNA profiling of degraded low copy number DNA samples, mtDNA PCR quality evaluation and DNA typing for 16 autosomal STR and 9 Y chromosomal STR loci were per-formed and the correlation between DNA quantities and PCR amplification efficiencies of the samples was analyzed. The DNA quantities assayed by the simple method suggested in the present study could be good indicator for mtDNA and STR analysis. As the allele drop-out was observed in less than 0.050ng DNA samples, at least 0.100ng of DNA is required to produce informative STR profiles. Also, STRs with less than 200bp amplification sizes produce efficient DNA profiles in most cases. Therefore, the develop-ment of mini-STRs with less than 200bp amplification sizes is expected to improve DNA typing in degraded low copy number DNA. Y-STRs are easy to detect allele drop-out or drop-in, and accordingly the efficiency test of Y-STRs as well as autosomal STRs for profiling of degraded low copy number DNA samples is thought to be important.