SWGDAM Staff Scientist Thermo Fisher Scientific Nov 17

法庭科学SWGDAM��流程 �昌 Staff Scientist | Thermo. Fisher Scientific Nov 17 th, 2016 The world leader in serving science

�程 • SWGDAM 介� • �夏TM白金SWGDAM �� • Yfiler. Plus. TM SWGDAM �� • ISO 18385 介� 2

SWGDAM主� Source: swgdam. org 3

关于SWGDAM • First meeting of Technical Working Group on DNA Analysis Methods (TWGDAM) was held in November 1988, with a subcommittee on restriction fragment length polymorphism (RFLP) DNA analysis. • TWGDAM continued to provide a level of direction to the forensic DNA community by issuing guidelines for DNA analysis, and the guidelines became de facto standards and were recognized by courts. • SWGDAM Chairman is appointed by the FBI Director, and members are selected by the Chairman from forensic DNA laboratories. • SWGDAM has multiple committees including Autosomal STR Committee, CODIS Committee, Rapid DNA Committee, Next Generation Sequencing Working Group, etc. • One of SWGDAM’s most important responsibilities is the recommendation of revisions to the FBI’s Quality Assurance Standards (QAS) for DNA Analysis. 4

SWGDAM 出版物 5

SWGDAM法医DNA分析方法��指� 3. 1 Genetic Markers 3. 6 Case Type Samples 3. 2 Species Specificity 3. 7 Population Studies 3. 3 Sensitivity 3. 8 Mixture Studies 3. 4 Stability 3. 9 PCR Based Studies 3. 5. Precision & Accuracy SWGDAM Validation Guidelines for Forensic DNA Analysis Methods approved in November 2012 www. swgdam. org 6

�夏白金物种特异性 – 非灵�� 200 rfu Bovine Chicken • • 7 SWGDAM 3. 2 The ability to detect genetic information from non-targeted species (e. g. , detection of microbial DNA in a human assay) should be determined. 700 rfu Pig Dog Rabbit Horse Rat Microbes Sheep Mouse NTC 200 rfu 10 ng non-primate DNA, or 105 copies of microbes (species commonly found in the oral cavity, listed in User Guide) were added to the system using 29 PCR cycles. Low peaks were detected with bovine, dog (like contamination, also found in the repeat), horse, mouse and pig.

�夏白金一次性成功通�率 SWGDAM 3. 4 The ability to obtain results from DNA recovered from biological samples deposited on various substrates and subjected to various environmental and chemical insults should be evaluated. • All sample types have ≥ 90% pass rate (PAT = 175 rfu) with 1 Proportion test. • The recommended rxn volume for HXP is 25 ul, some tests here were done with 10 ul reaction volume to challenge the system. 8

�本准在 3500 xl上的准确度 SWGDAM 3. 5 Precision and accuracy of the assay should be demonstrated. Accuracy is the degree of conformity of a measured quantity to its actual (true) value. Size deviation of 42 population samples showed deviation <0. 50 bp for alleles 9

����在不同种族的人群中的分布 SWGDAM 3. 7 The distribution of genetic markers in populations should be determined in relevant population groups. • Allele frequency for major Chinese population groups was studied. • Data of D 6 S 1043 was presented above, and the data with other markers was published in UG. 10

反��混物保��研究 Individual components Component concentrations • The Huaxia Platinum MM is robust to raw material change 11 SWGDAM 3. 9 The reaction conditions needed to provide the required degree of specificity and robustness should be determined. • Eight raw materials in master mix were varied up to 20%. • ICB was above 35% for most raw materials with 10% or even 20% variation. • Detail data including capability analysis presented next.

SWGDAM �Y-STR的指� • Y-STR typing is also used in lieu of autosomal typing for the detection of male DNA in mixtures that contain an overabundance of female DNA. Given that under certain conditions a male minor contributor in a mixture of female: male DNA may only be detectable by Y-STR typing, laboratories should pursue Y-STR analysis as the most appropriate means of detecting a male contributor(s) in some forensic samples Interpretation Guidelines for Y-Chromosome STR Typing approved in January 2014 www. swgdam. org This SWGDAM statement highlights the importance of male specificity of Y-STR kit with the male female mixtures commonly found with the sex assault cases. 12

提高Y-特异性的 作 • Primer mix • Design/screen primer pairs with minimal female crossreactivity • Master mix • Lower the Mg and Taq enzyme concentrations to minimize female cross-reactivity • Multiple rounds of DOE were performed to optimize the MM components • PCR condition • Increase the annealing/extension temperature to increase male specificity 13

Yfiler Plus 在男女混合DNA的表� Internal Product Spec: The kit will recover full profile with average 40% ICB when amplified with male: female mixtures of 1: 1000 • Yfiler Plus recovered full profile with 1: 1000 male: female mixture • No extra peaks with Yfiler Plus, while multiple extra peaks with PPY 23 and Yfiler 14

Yfiler Plus高�度女性DNA下的基� Internal Product Spec: The Yfiler Plus kit shall not amplify reproducible female DNA in the 1 ug range, without reproducible drop ins & artifacts within the read region. 200 rfu 68 bp 406 bp Blue 271 Outside the read region Red 139/144 Red grass Low female cross-reactivity (<175 PAT) within the read region are all documented in the UG. 15

3 微克女性DNA在三个盒子的比� PAT 3 ug of DNA from 10 different female donors were tested with the Yfiler Plus, PPY 23 and Yfiler kits. The highest reproducible female artifacts from each sample were plotted by kit. Yfiler Plus and Yfiler have significantly lower artifact peak heights than PPY 23. 16

新的法医DNA�准 – ISO 18385 Manufacturing Quality Commitment Meet ISO 18385 requirements for the manufacture of Forensic DNA Grade reagents Provide same level of manufacturing quality and QC testing across HID product lines Both Yfiler plus and Huaxia Platinum kits are ISO 18385 compliant 17