Genetic characterization of four dog breeds with Illumina CanineHD BeadChip

Dear Editor,Due to human’s cohabitation with domesticated animals,molecular analysis of animal DNA is increasingly being admitted as evidence in forensic investigations.In 2011,recommendations from the International Society of Forensic Genetics(ISFG)for non-human DNA analysis in forensic casework were published based on the successful model for human DNA[1].Among domesticated animals,canine DNA is perhaps the most often encountered and investigated in the forensic community[2–5].The US,Brazil and China are the top three countries in regards to ownership of canines.Canine DNA in the form of hair,saliva,blood,urine and feces is abundant in the domestic environment and consequently is often present on evidence collected during forensic investigations.A strong need for identity identification,parentage verification and breed recognition has become apparent within the forensic community.

Multi-locus identification of Psilocybe cubensis by high-resolution melting(HRM)

Hallucinogenic mushroom is a kind of toxic strain containing psychoactive tryptamine substances such as psilocybin,psilocin and ibotenic acid,etc.The mushrooms containing hallucinogenic components are various,widely distributed and lack of standard to define,which made a great challenge to identification.Traditional identification methods,such as morphology and toxicology analysis,showed shortcomings in old or processed samples,while the DNA-based identification of hallucinogenic mushrooms would allow to identify these samples due to the stability of DNA.In this paper,four primer sets are designed to target Psilocybe cubensis DNA for increasing resolution of present identification method,and the target markers include largest subunit of RNA polymerase II(marked as PC-R1),psilocybin-related phosphotransferase gene(marked as PC-PT),glyceraldehyde 3-phosphate dehydrogenase(marked as PC-3)and translation EF1α(marked as PC-EF).Real-time PCR with high-resolution melting(HRM)assay were used for the differentiation of the fragments amplified by these primer sets,which were tested for specificity,reproducibility,sensitivity,mixture analysis and multiplex PCR.It was shown that the melting temperatures of PC-R1,PC-PT,PC-3 and PC-EF of P.cubensis were(87.93±0.12)℃,(82.21±0.14)℃,(79.72±0.12)℃ and(80.11±0.19)℃ in our kinds of independent experiments.Significant HRM characteristic can be shown with a low concentration of 62.5pg/µL DNA sample,and P.cubensis could be detected in mixtures with Homo sapiens or Cannabis sativa.In summary,the method of HRM analysis can quickly and specifically distinguish P.cubensis from other species,which could be utilized for forensic science,medical diagnosis and drug trafficking cases.

Investigation on the genetic-inconsistent paternity cases using the MiSeq FGx system

Mutations might challenge the paternity index calculation in forensic identification.While many studies have focussed on the autosomal short tandem repeats(A-STR),the mutation status of sex chromosomes and single nucleotide polymorphism(SNP)remain blank.Next generation sequencing(NGS),known as high throughput and large sequence polymorphism,is a promising tool for forensic genetics.To describe the mutation landscapes in the paternity cases with genetic inconsistencies,a total of 63 parentage confirmed paternity cases contained at least one mismatched locus have been collected.The mutations were subsequently evaluated using Verogen’s MPSForenSeqTM DNASignature Kit and a microsatellite instability(MSI)detection kit.The result showed 98.41%(62/63)of the cases had no additional autosomal mutations even when the number of A-STRs increased to 27.As for the sex chromosomes,about 11.11%(7/63)of the cases exhibited either X-STR or Y-STR mutations.D2S1338,FGAand Penta Ewere the most frequent altered STRs,which suggested they might be the mutation hotspots.In addition,a male with sex chromosome abnormality was observed accidently,whose genotype might be 47,XXY,rather than MSI.Nearly 56.90%of the STR loci possessed isoalleles,which might result in higher STR polymorphisms.No Mendelian incompatibility was detected among the SNP markers,which indicated that SNP was a more reliable genetic marker in the genetic-inconsistent paternity cases.

Validation studies of the Para DNA^(■) Intelligence System with artificial evidence items

Short tandem repeat(STR)profiling is one of the mostly used systems for forensic applications.In certain circumstances,STR profiling is time-consuming and costly,which potentially leads to delays in criminal investigations.LGC(Laboratory of the Government Chemist,UK)Forensics has developed a robust STR profiling platform called the ParaDNAVR Intelligence Test System which can provide early tactical intelligence and aid investigators in making informed decisions on sample prioritization for detection.Here,we validated the ParaDNA^(■) intelligence test for its application in forensic cases using a range of mock evidence items following guidelines set by the Scientific Working Group on DNA Analysis Methods(SWGDAM).Specifically,we tested the sensitivity and accuracy of the ParaDNA intelligence test,as well as the success rates for detecting mock samples and for use in case scenarios.Our findings demonstrate that the ParaDNA intelligence test generates useful DNA profiles,especially for samples such as blood,saliva,and semen that contain ample DNA,indicating the benefits of including ParaDNA as a prior step in forensic STR profiling pipelines.

Validation of the Investigator 24plex QS Kit:a 6-dye multiplex PCR assay for forensic application in the Chinese Han population

The Investigator 24plex QS Kit(QIAGEN,Hilden,Germany)is a 6-dye fluorescent chemistry short tandem repeat(STR)polymerase chain reaction(PCR)amplification system that simultaneously amplifies 20 of the expanded Combined DNA Index System(CODIS)core STR loci,SE33,DYS391,and the standard sex-determining locus,amelogenin,as well as two special internal performance quality sensor controls(QS1 and QS2),which are included in the primer mix to check the PCR performance.This study was designed to be a pilot evaluation of this STR-PCR kit in a Chinese Han population regarding the PCR conditions,sensitivity,precision,accuracy,repeatability,reproducibility,and concordance;tolerance to PCR inhibitors;applicability to real“forensic-type”samples;species specificity;mixture,balance and stutter analyses,and utility in a population investigation.The exhaustive validation studies demonstrated that the Investigator 24plex QS system is accurate,sensitive and robust for STR genotyping.In addition,these genetic markers in the population data in our study indicated that they can also be useful for forensic identification and paternity testing in the Chinese Han population.

Forensic parameters of 41 Y-STR loci in Shandong Han individuals and comparison with 42 other populations

The Y-chromosomal short tandem repeat polymorphisms(Y-STRs)are the male-specific markers.The characteristics of paternal lineages make it a valuable tool for tracing familial relationships[1-3].Y-STR analysis has been widely used for identifying genealogical DNA testing and to identify missing persons,assess paternal relationships,and investigate sexual assault cases[4,5].

Genetic polymorphisms of 21 STR loci of Goldeneye^(TM) DNA ID 22NC kit in five ethnic groups of China

Dear Editor,Short tandem repeats(STRs),polymorphic DNA regions with a variable number of repeated units(2–6 base pairs),are attractive to forensic applications such as human identification and parentage testing[1].Nowadays,most of the commercial STR kits are designed based on STRs from the combined DNA index system(CODIS),European Standard Set(ESS),expanded CODIS,and extended ESS[2].In this study,we evaluated 21 STRs from GoldeneyeTM DNA ID 22NC kit(PeopleSpot Inc.,Beijing,China),which including 20 polymorphic non-CODIS STR loci(i.e.D1S1656,D2S441,D3S1744,D3S3045,D4S2366,D5S2500,D6S477,D7S1517,D7S3048,D8S1132,D10S1248,D10S1435,D11S2368,D13S325,D14S608,D15S659,D17S1290,D18S535,D19S253,D22GATA198B05)and a CODIS STR locus(D3S1358),in five ethnic groups(i.e.Eastern Han,Ningxia Hui,Xinjiang Uygur,Xizang Tibetan,and Inner Mongolia Mongolian)of China.The forensic genetic investigation of above loci may provide more genetic information in complex kinship testing and population studies[3,4].