scholarly journals Assessment of autosomal and male DNA extracted from casework samples using Casework Direct Kit, Custom and Maxwell 16 System DNA IQ Casework Pro Kit for autosomal-STR and Y-STR profiling

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Hashom Mohd Hakim ◽  
Hussein Omar Khan ◽  
Siti Afifah Ismail ◽  
Shahrizad Ayob ◽  
Japareng Lalung ◽  
...  
Keyword(s):  
Tandem Repeats ◽  
Pcr Amplification ◽  
Dna Profiling ◽  
Nucleic Acid Content ◽  
Str Profiling ◽  
Autosomal Str ◽  
Y Chromosomes ◽  
Y Chromosome Str ◽  
Equal Capacity ◽  
Dna Template

Abstract Short repetitive regions in autosomal and Y chromosomes known as short tandem repeats (STRs) are currently used for DNA profiling in crime investigations. However, DNA profiling requires a sufficient quality and quantity of DNA template, which is often not obtained from trace evidence or degraded biological samples collected at the scene of a crime. Here, we assessed autosomal and male DNA components extracted from crime scene and mock casework samples using the Casework Direct Kit, Custom and compared the results against those obtained by extraction of matching samples using well-established Maxwell 16 System DNA IQ Casework Pro Kit. The quantity and quality of extracted DNA obtained using both Casework Direct Kit, Custom and Maxwell 16 System DNA IQ Casework Pro Kit were analyzed using PowerQuant Systems followed by autosomal and Y-chromosome STR profiling using GlobalFiler Express PCR Amplification Kit and PowerPlex Y23 System, respectively. Our results showed that the Casework Direct Kit and Maxwell 16 DNA IQ Casework Pro Kit have more or less equal capacity to extract inhibitor free DNA, but that the latter produces slightly better quality and more DNA template and subsequently higher numbers of STR allele calls for autosomal and Y-STR analyses. Nonetheless, the Casework Direct Kit, Custom is the quicker and cheaper option for extraction of good, clean DNA from high content material and might best be used for extraction of reference samples. Such reference DNA samples typically come from buccal swabs or freshly drawn blood. So, in general, they can confidently be expected to have a high nucleic acid content and to be inhibitor-free.

scholarly journals Assessment of QIAGEN™ Investigator® 24plex GO! kit workflow for autosomal STR profiling of forensic reference samples

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Hashom Mohd Hakim ◽  
Hussein Omar Khan ◽  
Siti Afifah Ismail ◽  
Nurul Hazirah Mat Lazim ◽  
Japareng Lalung ◽  
...  
Keyword(s):  
Pcr Amplification ◽  
Dna Profiling ◽  
Buccal Swab ◽  
Forensic Dna ◽  
Str Typing ◽  
Str Analysis ◽  
Dna Database ◽  
Autosomal Str ◽  
Dna Template ◽  
Reference Samples

Abstract Background DNA profiling has proven to be a valuable technique for identification of individuals in crime. Currently, the technique targets several short tandem repeat (STR) regions in human genome. However, increasing number of samples submitted for STR analysis may lead to delays due to the limited number of experienced analysts who might be available at any given moment and the time taken to complete lengthy DNA profiling procedures. This study was conducted to test the specificity, repeatability, reproducibility and robustness of Investigator® 24plex GO! kit for genotyping of reference samples submitted to the Royal Malaysian Police Forensic DNA Laboratory for DNA database. Material and methods In this study, Investigator® 24plex GO! kit was used to directly amplify STR loci from buccal swab cell of reference samples that had previously been STR typed using GlobalFiler™ Express kit. Capillary electrophoresis was carried out on a 3500xL Genetic Analyser using POP-4® Polymer. Amplified products were assigned to particular STR alleles using the GeneMapper ID-X version 1.4 software. Results Our study shows that STR profiles generated using Investigator® 24plex GO! gave concordance results with those previously obtained using the GlobalFiler™ Express kit. In addition, quality sensors included in the kit are of particular importance for determining the effectiveness of the PCR reaction and help to indicate the nature and quantity of DNA template for PCR amplification. Conclusion The Investigator® 24plex GO! kit is reliable for STR typing of reference samples.

scholarly journals Sperm Cell Capture Based on ABH Antigen Differences to Separate Two Men in Mixed Seminal Stains

2021 ◽  
Vol 2021 ◽  
pp. 1-5
Author(s):  
Mao-ling Sun ◽  
Ji-long Zheng ◽  
Bao-jie Wang ◽  
Jun Yao
Keyword(s):  
Sperm Cell ◽  
Tandem Repeats ◽  
Pcr Amplification ◽  
Personal Identification ◽  
Blood Type ◽  
Cell Capture ◽  
Str Typing ◽  
Autosomal Str ◽  
Unrelated Donors ◽  
Seminal Stains

Personal identification of two individuals in mixed semen samples in forensic DNA testing in general usually involves analysis using autosomal and Y chromosome short tandem repeats (STRs). Results may exclude unrelated donors but cannot identify individuals. In this study, sperm cell capture based on ABH antigen differences was used to obtain the cells with the single ABO blood type. Immunohistochemical staining using labeled anti-A, anti-B, and anti-H antibodies and the laser microdissection system can be used to enrich sperm with different ABO types in mixed seminal stains from two individuals. Then, PCR amplification and capillary electrophoresis were performed to genotype the STR loci. To some extent, after sperm cell capture based on ABH antigen differences, autosomal STR typing using enriched single blood group cells can be utilized to partially identify different individuals in a mixed seminal stain sample from two individuals.

scholarly journals Validation Study of 6 Dye-Based SF25TM PCR Amplification Kit

2021 ◽  
Author(s):  
Pankaj Shrivastava ◽  
R. K. Kumawat ◽  
Pushpesh Kushwaha ◽  
Akshay Kumar ◽  
Manisha Rana ◽  
...  
Keyword(s):  
Dna Analysis ◽  
Pcr Amplification ◽  
Turnaround Time ◽  
Fluorescent Dye ◽  
Forensic Dna ◽  
Allelic Ladder ◽  
Forensic Dna Analysis ◽  
Inhibitor Tolerance ◽  
Autosomal Str ◽  
Dna Template

Abstract Ever since the inception of STR-based forensic DNA analysis for human Identification and DNA profiling, the approach developed continuously. Most of these developments are pertaining to achieve the best polymorphic loci set for analysis, high inhibitor tolerance capability, increase in sensitivity, reduction in turnaround time, and cost-effectiveness. On the similar line, the SF25TM PCR amplification kit, a 6 fluorescent dye-chemistry based autosomal STR kit was developed, which analyses 25 STR loci viz., FAM fluorescent dye-labelled D3S1358, D13S317, D7S820, D16S539, D1S1656, and Penta E; HEX fluorescent dye-labelled TPOX, TH01, D2S1338, CSF1PO, and Penta D; SUM fluorescent dye-labeled D19S433, vWA, D21S11, D18S51, and D6S1043; LYN fluorescent dye-labelled Amelogenin, D8S1179, D5S818, D12S391, and FGA; PUR fluorescent dye-labelled D22S1045, Y indel; D2S441 and D10S1248, The developmental validation of the SF25 PCR Amplification reagents were carried out in this study to analyze its specificity, sensitivity, and concordance. This multiplex kit showed higher sensitivity with 125pg of DNA template generating a complete profile (with 100% allele call). Out of 83 samples, 10 samples showed aberrant triallelic pattern/aberrant heterozygous pattern at D16S539 locus. The possible discrepant homozygous condition at locus D1S1656 might be due to the invasion of alleles at D1S1656 locus present adjacent to D16S539 locus. Such invasion resulted due to the shortening of the allelic bin at the D1S1656 locus. The allelic ladder of the SF25TM PCR amplification kit reveals the allelic range of D1S1656 from 11 to 19.3 which can be interpreted according to the casework as deemed by the end-users for their samples.

scholarly journals ISOLASI FRAGMEN GEN PENYANDI PUTRESIN N-METILTRANSFERASE DAN QUINOLINAT FOSFORIBOSILTRANSFERASE ASAL TEMBAKAU LOKAL TEMANGGUNG (Nicotiana tabacum)

2020 ◽  
Vol 17 (3) ◽  
pp. 109 ◽  
Author(s):  
SESANTI BASUKI ◽  
NURHAJATI AA MATTJIK ◽  
SUWARSO SUWARSO ◽  
DESTA WIRNAS ◽  
SUDARSONO SUDARSONO
Keyword(s):  
Dna Sequences ◽  
Pcr Amplification ◽  
Data Bank ◽  
Degenerate Primer ◽  
Gene Encoding ◽  
Methyl Transferase ◽  
Dna Database ◽  
Genes Encoding ◽  
Tobacco Cultivar ◽  
Dna Template

<p>ABSTRAK</p><p>Upaya untuk menurunkan kandungan nikotin merupakan salah satuprioritas utama penelitian tembakau. Nikotin adalah senyawa alkaloidutama berpotensi dikonversi menjadi senyawa nor-nikotin yang bersifatkarsinogen. Gen PMT sebagai penyandi enzim putresin n-metiltransferase(PMT) dan gen QPT - penyandi enzim quinolinat fosforibosiltransferase(QPT) merupakan dua gen kunci yang berperan penting pada proses bio-sintesis nikotin. Penelitian ini bertujuan untuk mengisolasi potongan genPMT dan QPT asal tembakau lokal Indonesia, mengkarakterisasi danmenganalisis runutan DNA-nya. Tahapan penelitian dimulai dengan me-rancang primer degenerate berdasarkan informasi yang ada di pangkalandata Bank Gen NCBI (National Centre for Biotechnology Information),mengamplifikasi PCR menggunakan templat DNA genomik tembakaulokal cv. Sindoro1, mengklon potongan DNA hasil PCR dan menentukanrunutan DNA-nya. Hasil penelitian menunjukkan dari dua belas pasangprimer degenerate yang dirancang, hanya dua pasang primer yang meng-hasilkan potongan DNA hasil amplifikasi PCR, yaitu pasangan primerPMt-7 (F &amp; R) untuk gen PMT dan primer QPt-3 (F &amp; R) untuk gen QPT.Setelah dilakukan penentuan runutan DNA-nya, amplikon yang didapatdari hasil PCR dengan pasangan primer PMt-7 sebesar 1418 bp, sedangkanuntuk primer QPt-3 sebesar 205 bp. Runutan DNA gen PMT dan gen QPTasal tembakau lokal cv. Sindoro1 mempunyai tingkat kesamaan yang ting-gi dengan gen PMT dan gen QPT asal tembakau lainnya yang ada dipangkalan data Bank Gen NCBI.</p><p>Kata kunci : Gen PMT, gen QPT, lintasan biosintesis nikotin, perunutanDNA, amplifikasi PCR, primer degenerate</p><p>ABSTRACT</p><p>Isolation of Genes encoding Putrescine N-Methyl-transferase and Quinolinat Phosphoribosyl transferasederived from Temanggung Tobacco Cultivar (Nicotianatabacum)</p><p>Reduction of nicotine content is one of the major objective intobacco research. Nicotine is the main alcaloid compound that potentiallycould be converted into a carcinogenic compound (nor-nicotine). The PMTgene encoding putrescine N-methyl transferase (PMT) and the QPT gene -encoding quinolinate phosphoribosyl transferase (QPT) are the two keyenzymes involved in nicotine biosynthesis. The objectives of this researchwere to isolate PMT and QPT gene fragments originated from Indonesianlocal tobacco, to characterize, and to analyze their DNA sequences. Theresearch activities included: degenerate primer design based oninformation available in the GenBank DNA Database NCBI (NationalCentre for Biotechnology Information), PCR amplification usingdegenerate primer and genomic DNA template of a local tobacco cv.Sindoro1, clone the PCR amplified products, and determine their DNAnucleotide sequences. Results of the experiment indicated that from 12degenerate primer pairs synthesized, only two were able to yield positivePCR amplified products. These primer pairs were PMt-7 (F &amp; R primers)for PMT and QPt-3 (F &amp; R primers) for QPT. After DNA sequencing, theamplified DNA product amplified using PMt-7 degenerate primer pairswere 1418 bp, while that using QPt-3 primer pairs were only 205 bp.Nucleotide sequences of PMT or QPT gene fragments originated fromlocal tobacco cv. Sindoro1 showed a high nucleotide sequences identity ascompared to that of the respective genes from other tobacco species thatwere available in the GenBank DNA Database NCBI.</p><p>Key words: PMT gene, QPT gene, nicotine biosynthetic pathways, DNAsequencing, PCR amplification, degenerate primer</p>

scholarly journals Identification of a member of a DNA-dependent ATPase family that causes interference with silencing.

1997 ◽  
Vol 17 (9) ◽  
pp. 5461-5472 ◽  
Author(s):  
Z Zhang ◽  
A R Buchman
Keyword(s):  
Mating Type ◽  
Tandem Repeats ◽  
Specific Interaction ◽  
Cis Acting ◽  
Cellular Processes ◽  
Protein Dna Interactions ◽  
Mating Type Switching ◽  
Silencer Elements ◽  
Dna Template ◽  
Knockout Mutation

DNA in eukaryotic cells is packed in tandem repeats of nucleosomes or higher-order chromatin structures, which present obstacles to many cellular processes that require protein-DNA interactions, such as transcription, DNA repair, and recombination. To find proteins that are involved in increasing the accessibility of specific DNA regions in yeast, we used a genetic approach that exploited transcriptional silencing normally occurring at HML and HMR loci. The silencing is mediated by cis-acting silencer elements and is thought to require the formation of a special chromatin structure that prevents accessibility to the silenced DNA. A previously uncharacterized gene, termed DIS1, was isolated from a screen for genes that interfere with silencing when overexpressed. DIS1 encodes a protein with conserved motifs that are present in a family of DNA-dependent ATPases, the SWI2/SNF2-like proteins. Overproduction of N-terminal half of DIS1 protein interfered specifically with ectopic silencing used in the screen as well as HMR E silencing. Two-hybrid studies revealed a specific interaction between the N terminus of DIS1 and the C-terminal half of SIR4, a protein essential for silencing. Cells with a dis1 knockout mutation had significantly lower mating-type switching rate. These results suggest that DIS1 may contribute to making the silenced DNA template at HM loci more accessible during the mating-type switching process.

Genetic diversity of Mycoplasma hyopneumoniae isolates from conventional farrow-to-finish pig farms in Serbia

2010 ◽  
Vol 58 (3) ◽  
pp. 297-308 ◽  
Author(s):  
Bozidar Savic ◽  
Vojin Ivetic ◽  
Vesna Milicevic ◽  
Ivan Pavlovic ◽  
Milenko Zutic ◽  
...  
Keyword(s):  
Tandem Repeats ◽  
Genetic Relatedness ◽  
Pcr Amplification ◽  
Mycoplasma Hyopneumoniae ◽  
Variable Number ◽  
Repeat Motif ◽  
Typing Method ◽  
Pig Farms ◽  
Different Ages ◽  
Swine Population

Mycoplasma hyopneumoniaeis a primary agent associated with mycoplasma pneumonia and the porcine respiratory disease complex (PRDC). Various reports have indicated that different strains ofM. hyopneumoniaeare circulating in the swine population. Lysates from lung swabs from naturally infected pigs of different ages were tested according to a new variable number of tandem repeats (VNTR) genetic typing method based on the polyserine repeat motif of the P146 lipoproteoadhesin, which can be applied directly on clinical material without isolation ofM. hyopneumoniae. The aim was to determine the diversity ofM. hyopneumoniaeisolates from conventional farrow-to-finish pig farms located in different geographical areas of Serbia. PCR amplification was carried out usingM. hyopneumoniae-specific designed, conserved primers (p146MH — L and p146MH — R) flanking the region encoding the repeat motif, followed by sequencing and cluster analysis. Five groups ofM. hyopneumoniaewith thirteen to twenty-four serine repeats were observed. Analysis of three samples from each farm indicated that the specific isolate is ubiquitous in pigs of different ages. Furthermore, seven clusters were observed within 27 tested samples. The results indicated a considerable diversity amongM. hyopneumoniaefield isolates in the swine population from conventional farrow-to-finish farms in Serbia and suggest close genetic relatedness of the corresponding isolates.

Characterization of the Gyrodactylus salaris Malmberg, 1957 (Platyhelminthes: Monogenea) ribosomal intergenic spacer (IGS) DNA

Parasitology ◽  
2000 ◽  
Vol 121 (5) ◽  
pp. 555-563 ◽  
Author(s):  
C. M. COLLINS ◽  
C. O. CUNNINGHAM
Keyword(s):  
Ribosomal Rna ◽  
Tandem Repeats ◽  
Gene Array ◽  
Pcr Amplification ◽  
Intergenic Spacer ◽  
Sequence Motifs ◽  
Ribosomal Rna Gene ◽  
Intergenic Spacers ◽  
Transcription Start Sites ◽  
Pcr Products

The intergenic spacer of the ribosomal RNA gene array from the monogenean Gyrodactylus salaris was isolated using PCR amplification. PCR products were cloned and sequenced. Three different fragments of 0·63, 1·0 and 2·62 kb, were consistently obtained. These showed homology at the 5′ and 3′ termini but differed in their overall size and intervening sequence. The 5′ end showed homology to various 28S ribosomal RNA gene sequences, suggesting that this represented the 3′ terminus of the G. salaris 28S ribosomal RNA gene. A number of features common to other eukaryotic intergenic spacers were found in the longest sequence, including A + T rich sequences, palindromic sequences and tandemly repeated elements. Two regions of 23 bp sequences arranged in non-identical tandem repeats were identified. There were 9 repeats in both regions, separated by 81 bp of non-repetitive sequence. The repeat units from the two regions shared some similarity at their 3′ ends. The G. salaris intergenic spacer sequence was examined for sequence motifs involved in the transcription of the ribosomal RNA genes in other species. Several regions with homology to transcription start sites were identified.

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