Effective DNA Extraction Research Articles

DNA Barcoding Unveils Novel Discoveries in Authenticating High-Value Snow Lotus Seed Food Products.

Snow Lotus Seed (SLS), esteemed for its nutritional and market value, faces challenges of authentication due to the absence of appropriate testing standards and methods. This results in frequent adulteration of SLS sourced from Gleditsia sinensis (G. sinensis) with other plant seeds endosperm. Traditional chloroplast DNA barcoding methods are inadequate for species identification due to the absence of chloroplasts in G. sinensis seeds endosperm. In this study, the homology of 11 ITS genes among 6 common Gleditsia species was analyzed. Universal primers suitable for these species were designed and screened. A DNA barcoding method for distinguishing SLS species was developed using Sanger sequencing technology, leveraging existing GenBank and Barcode of Life Data System (BOLD) databases. Optimized sample pretreatment facilitated effective DNA extraction from phytopolysaccharide-rich SLS. Through testing of commercial SLS products, the species origin has been successfully identified. Additionally, a novel instance of food fraud was uncovered, where the Caesalpinia spinosa endosperm was used to counterfeit SLS for the first time. The study established that the developed DNA barcoding method is effective for authenticating SLS species. It is of great significance for combating food fraud related to SLS, ensuring food safety, and promoting the healthy development of the SLS industry.

An effective DNA extraction method from Cajanus cajan seeds suitable for PCR analysis

La presencia de impurezas extraídas conjuntamente en las semillas puede dificultar la extracción de ADN, ya que estos contaminantes pueden interferir con la amplificación por PCR, ambos procesos son pasos importantes en los programas continuos de reproducción y selección. En este estudio se evalúa un nuevo protocolo como alternativa para extraer ADN de los ápices de la radícula embrionaria de semillas de guandú que producen ADN fácilmente adecuado para la PCR. El protocolo propuesto para la extracción de ADN en semillas de guandú muestra un gran potencial como método alternativo, ya que es económico, sencillo y efectivo para la amplificación por PCR. También tiene la ventaja de que no requiere reactivos costosos o peligrosos y puede ampliarse fácilmente, lo que demuestra su potencial como un recurso valioso para los científicos que estudian la genética y el mejoramiento del guandú.

An innovative optimized protocol for high-quality genomic DNA extraction from recalcitrant Shea tree (Vitellaria paradoxa, C.F. Gaertn) plant and its suitability for downstream applications.

It is not always easy to find a universal protocol for the extraction of genomic DNA (gDNA) from plants. Extraction of gDNA from plants such as shea with a lot of polysaccharides in their leaves is done in two steps: a first step to remove the polysaccharides and a second step for the extraction of the gDNA. In this work, we designed a protocol for extracting high-quality gDNA from shea tree and demonstrate its suitability for downstream molecular applications. Fifty milligrams of leaf and root tissues were used to test the efficiency of our protocol. The quantity of gDNA was measured with the NanoDrop spectrometer and the quality was checked on agarose gel. Its suitability for use in downstream applications was tested with restriction enzymes, SSRs and RAPD polymerase chain reactions and Sanger sequencing. The average yield of gDNA was 5.17; 3.96; 2.71 and 2.41µg for dry leaves, dry roots, fresh leaves and fresh roots respectively per 100mg of tissue. Variance analysis of the yield showed significant difference between all tissue types. Leaf gDNA quality was better compared to root gDNA at the absorbance ratio A260/280 and A260/230. The minimum amplifiable concentration of leaf gDNA was 1pg/µl while root gDNA remained amplifiable at 10pg/µl. Genomic DNA obtained was also suitable for sequencing. This protocol provides an efficient, convenient and cost effective DNA extraction method suitable for use in various vitellaria paradoxa genomic studies.

An improved and effective DNA extraction protocol for Pyracantha crenulata with optimal PCR reliability

Abstract There are many challenges involved in obtaining high-quality DNA from tissues/cells, especially when using plants as the raw material. Three different DNA extraction protocols were employed in order to isolate high quality of genomic DNA from Pyracantha crenulata leaves. P. crenulata is a complex, versatile, and evergreen shrub species in Rosaceae family which is ecologically, economically and culturally important. This species produces valuable antioxidants like polyphenols, polysaccharides and secondary metabolites which interfere with DNA extraction thus making molecular marker based studies difficult in this species. This study aimed to develop a simple, rapid, cost-effective and highly efficient protocol for P. crenulata leaves rich in salts and phenols. The results demonstrated that modified CTAB (double buffer) protocol is very effective in overcoming the challenges that could impede next-generation sequencing analysis. The yield of the extracted DNA was excellent ranging from 368 to 410 ng/μL DNA with A260/280 ratio ranging from 1.80 to 1.84. The extracted DNA was amenable to PCR amplification making it suitable for DNA-based molecular marker studies. None procedures have been published for the purpose up this species, therefore, double buffer based extraction protocol is reported for the first time in P. crenulata and might prove useful in other species of family Rosa-ceae. This study describes an effective DNA isolation method that is appropriate for frequent population genetic screening. In conclusion, the double buffer protocol showed the most efficient and effective approach for extracting DNA from phenol rich, fresh, mature, and dry leaves.

Comparison of DNA extraction methods from soil samples

This paper reviews and compares various methods of extracting total DNA from soil samples using laboratory materials and reagents. The methods of DNA extraction were analyzed in different modifications and combinations with each other and with pretreatment of soil samples. It is shown that for effective DNA extraction, pretreatment of samples with calcium salts is necessary, in particular, the addition of CaCl2 to the extraction buffer to precipitate humic acids. DNA was extracted from the soil by all the methods, but the greatest amount of DNA was observed using a technique based on ethylenediaminetetetraacetic acid (EDTA), hexadecyltrimethylammonium bromide (CTAB), sodium dodecyl sulphate (SDS), and chloroform.

Effective DNA extraction method for metagenomic analysis of rhizosphere bacteria from mangrove sediments

Mangrove, Rhizophora mucronata, grows in the intertidal area, which contains much organic matter and varying salinity. The organic matter content is influenced by the bacterial community that inhabits the ecosystem, but information regarding the bacterial community, especially in the mangrove root system, is not widely available. There are several challenges in completing this information, one of which is that the method used is still in a conventional form. Developments in environmental DNA analysis can support and complement this information. However, this method must be optimized because the organic matter content and salt variations affect the extraction results. Thus, this study aimed to determine the optimal approach for extracting bacterial DNA from mangrove sediments. The analysis used two methodologies: manual DNA extraction techniques based on buffer modification and DNA extraction kits. There were four different treatments, namely the soil DNA isolation plus kit (M1), the fecal / soil microbial quick-DNA miniprep kit (M2), glass powder with charcoal (M3), and glass powder with skimmed milk (M4). DNA samples were obtained from each method and assessed for concentration and purity using a nanodrop. In addition, the resulting DNA's quality was analyzed using 1.5% agarose. The results obtained were in the M2 treatment, which showed optimal results compared to the others. M2 uses a bead-based beating and spin column method to achieve optimal DNA concentration through high molecular weight. The DNA obtained was also protein-free, and several samples were contaminated with humic acid, namely KL.S1, KL.S4, and T7.S4.Keywords:Bacteria 16SBead beatingDNA ExtractionSedimentSpin column

High-molecular-weight DNA extraction for long-read sequencing of plant genomes: An optimization of standard methods.

Developing an effective and easy-to-use high-molecular-weight (HMW) DNA extraction method is essential for genomic research, especially in the era of third-generation sequencing. To efficiently use technologies capable of generating long-read sequences, it is important to maximize both the length and purity of the extracted DNA; however, this is frequently difficult to achieve with plant samples. We present a HMW DNA extraction method that combines (1) a nuclei extraction method followed by (2) a traditional cetyltrimethylammonium bromide (CTAB) DNA extraction method for plants with optimized extraction conditions that influence HMW DNA recovery. Our protocol produced DNA fragments (percentage of fragments >20 kbp) that were, on average, ca. five times longer than those obtained using a commercial kit, and contaminants were removed more effectively. This effective HMW DNA extraction protocol can be used as a standard protocol for a diverse array of taxa, which will enhance plant genomic research.

Evaluation and comparison of three methоds оf DNA extractiоn frоm Kazakh horse of the type Zhabe

Horse breeding is one of the main directions in the livestock industry of Kazakhstan. Horse breeding is an economically important part of breeding, as horses have a high productive value. The purpose of this article was to determine the most optimal from the point of view of economic efficiency of the scientific DNA isolation method for genotyping of the Kazakh horse. In this study, we performed the first step (DNA extraction) in genotyping the DNA of the Kazakh horse to further determine the signs of growth, meat and dairy productivity. Nowadays, new generation sequencing technologies have made scientifi c and technological progress in research. Obtaining highly concentrated and non-contaminated DNA is the main stage of successful analysis. For further genetic studies, a concentration of 50 ng/ml is required, which is considered minimal. In the study, three different DNA extraction methods from tissue were subjected to comparative analysis in order to evaluate and identify the most effective DNA extraction method from horse ear tips. Real-time PCR amplification based on gel electrophoresis and spectrophotometric measurements (Nano Drop) were used to evaluate the isolated DNA's quality and quantity. In addition, energy consumption, time, as well as the cost of analysis were evaluated. According to the obtained results, extraction using the commercial kit protocol was simple to use, effective, but relatively expensive. The phenol-chloroform and CTAB methods are the same in terms of DNA quality, but given the danger of the phenol component used in the phenol-chloroform method, the CTAB method turned out to be the most acceptable for genotyping, because it is safe, not expensive and effective.

DNA Extraction of Bone Through Demineralization.

In the field of forensic science, the DNA extraction of bone is utilized in investigations involving mass disasters, unidentified remains, and missing persons. However, bone samples can be challenging samples due to their exposure to extreme environmental conditions over long periods of time. The use of an effective DNA extraction method to properly isolate and purify the DNA is essential for bone samples. This chapter describes the DNA extraction of bone samples through a total demineralization protocol, which aims to entirely dissolve the bone matrix in order to access the DNA molecules.

Comparative Study of DNA Extraction Methods for the PCR Detection of Intestinal Parasites in Human Stool Samples.

Stool samples typically contain PCR inhibitors; however, helminths are difficult to lyse and can cause false-negative PCR results. We assessed the effective methods for extracting DNA from different kinds of intestinal parasites. We compared the most common DNA extraction methods from stool samples, including the phenol-chloroform technique with or without a bead-beating step (P and PB), a QIAamp Fast DNA Stool Mini Kit (Q), and a QIAamp PowerFecal Pro DNA Kit (QB). Genomic DNA was extracted from 85 stool samples collected from patients infected with Blastocystis sp., Ascaris lumbricoides, Trichuris trichiura, hookworm, and Strongyloides stercoralis. DNA quantity and DNA quality were evaluated via spectrophotometry, and DNA integrity was assessed by PCR. We found that P and PB provided higher DNA yields (~4 times) than when using Q and QB. However, P showed the lowest detection rate of PCR (8.2%), wherein only S. stercoralis (7 out of 20 samples) was detected. QB showed the highest detection rate of PCR (61.2%). After plasmid spikes, only 5 samples by QB were negative while 60 samples by P were still negative. Remarkably, QB could extract DNA from all the groups of parasites that we tested. These results indicate that QB is the most effective DNA extraction method for the diagnosis and monitoring of intestinal parasites via PCR.

Modified methods obtain high-quality DNA and RNA from anaerobic activated sludge at a wide range of temperatures

Anaerobic activated sludge is rich in humic substances and water, leading to significant differences in the stability of metagenomic DNA and metatranscriptomic RNA. Thus, it is of great difficulty to exact high-quality and high-yield DNA and RNA from them, especially those cultured at a wide range of temperatures. Here, we established fast and effective DNA and RNA extraction methods based on current commercial kits. The modified methods combined liquid nitrogen grinding with kits, achieving notable improvements in concentrations, yields, purity and integrities for both DNA and RNA. The ratios of OD260/280 of the metagenomic DNA were between 1.81 ± 0.03 and 1.83 ± 0.02, while OD260/280 and OD260/230 of the metatranscriptomic RNA ranged from 1.96 ± 0.01 to 2.13 ± 0.03 and from 1.94 ± 0.02 to 2.30 ± 0.03 respectively. Metagenomic DNA and metatranscriptomic RNA obtained by the modified methods perfectly met the requirements of second- and third-generation sequencing, providing valuable reference for extracting high-quality metagenomic DNA and metatranscriptomic RNA from environmental samples of high water content and humic substances under temperatures ranging from 18 °C to 52 °C.

A survey of the effects of common illicit drugs on forensic DNA analysis

Profiling of DNA associated with illicit drug packages and paraphernalia is a common investigative tool. In addition, research is being conducted regarding the analysis of trace DNA present within illicit drugs and on capsules. The application of trace DNA analysis to illicit drugs has the potential to identify individuals involved in their manufacture and distribution. However, the inhibitory effects of illicit drugs and related compounds on downstream DNA analysis has not yet been investigated. If drug-induced polymerase chain reaction (PCR) inhibition occurs, the quality or informativeness of the resultant DNA profile may be impacted. In this study, the effects of a range of drugs, diluents, adulterants, and synthetic precursors on both quantitative PCR (qPCR) data and short tandem repeat (STR) DNA profiling results were examined. Twenty-two compounds representative of drug compounds and adulterants which may be encountered in drug seizures were spiked with 1 ng/μL and 0.05 ng/μL of control DNA and underwent DNA quantification using Quantifiler™ Trio. A subset of 13 compounds, including the majority that indicated potential inhibition in Quantifiler™ Trio, underwent STR profiling with VeriFiler™ Plus to determine if inhibition also occurred at this stage. The effect of diluting the DNA extract on the extent of inhibition of STR profiling was also investigated. Internal PCR controls within the qPCR were not a reliable indicator of inhibition, although suppression of the short and long autosomal fragments was observed in the presence of many compounds, and four compounds gave inconclusive results. STR internal quality controls indicated inhibition in 5 of the 13 compounds, however, profiles were affected by the presence of 11 of the 13 compounds in various ways such as a decreased average relative fluorescence units (RFU), drop out of certain alleles (some based on allele size range of locus) leading to a decreased likelihood ratio (LR), an increase in the proportion of stutter peaks and the presence of split or shoulder peaks. All profiles improved following a dilution of the compound in the PCR and allowing the generation of LR values in excess of 1 × 1025, indicating inhibition occurred rather than DNA degradation. The data obtained show that removal of some of these compounds is required through an effective DNA extraction process for successful downstream trace DNA profiling. Upon successful PCR, the resultant DNA profiles provide the opportunity for opening new investigative avenues for law enforcement agencies.

DNA EXTRACTION TECHNIQUES FOR SOME WILD SPECIES FROM UZBEKISTAN

Although DNA extraction is already a routine procedure and protocols have been developed for a large number of plant species, various plant tissues and organs, solving the problem of extracting pure undegraded DNA from plant is the main step in molecular approaches for study plant organisms. The extraction of high-quality DNA from plant materials is an important factor for DNA barcoding. The effective DNA extraction protocol for samples with high content of admixture, which prevents the restriction reaction and the PCR, or degraded DNA should be established for each case. This work presents the comparison of DNA isolation techniques from silica gel dried and herbarium samples of some wild species collected in Uzbekistan. In this work, we optimized the technique for isolating the total DNA, on the examples of 16 representatives of Salvia, Dracocephalum, Phlomoides, Tulipa, Iris, Allium genera from Uzbekistan. The results of a comparative analysis of three methods of DNA extraction from leaf tissues of wild plants are represented in this article. DNA amount and purity degrees were measured using spectrophotometer. The efficiency of the DNA extraction method was assessed according to two criteria: the quantitative parameters -DNA concentration (ng/μl), and the qualitative parameters - DNA purity (nm). The DNA concentration variate from 758.1 ng/μl obtained with the CTAB method to 13.89 ng/μl obtained with commercial column extraction kits. λ = 260/280 ratio is range from 1.7 to 1.98 nm. Additionally, PCR amplification was realized to reflect the validation of the DNA isolation protocol, using specific primers of four chloroplast (rbcL, matK, psbA, trnL-F) and one nuclear (ITS) plant DNA barcode regions. The sequence length was 300–800 bp appropriately.

Postsynthetic Modification of the Magnetic Zirconium-Organic Framework for Efficient and Rapid Solid-Phase Extraction of DNA.

In recent years, several approaches have been applied to modify metal-organic frameworks (MOFs) owing to their excellent structural tunability such as higher extraction efficiency than that of primitive crystals. Herein, Zr-based MOFs (UiO-66-NH2) with a suitable size modulated by acetic acid were successfully synthesized for effective DNA extraction. The bonding conformations and adsorption mechanism indicated a high affinity between UiO-66-NH2 and the DNA molecules. Furthermore, Fe3O4 nanoparticles were immobilized on the UiO-66-NH2 surface to allow MOFs with magnetism. The magnetic zirconium-organic framework (MZMOF) retained the intact structure of MOFs and simplified subsequent extraction operations. In the DNA recovery investigation, MZMOF showed high recovery efficiency for both short-stranded DNA (90.4%) and pseudovirus DNA (95.1%). In addition, it showed superior DNA extraction efficiency from plasma (57.6%) and swab preservation solution (86.5%). The prepared MZMOF was employed for highly specific extraction of viral DNA and cfDNA from samples. To further simplify the extraction process, MZMOF was applied to immiscible phase filtration assisted by a surface tension (IFAST) chip for facilitating rapid DNA extraction with sensitive point-of-care testing. The developed MZMOF-based extraction method has significant potential for increasing the demand for rapid and efficient nucleic acid extraction.

Optimized DNA-based identification of Toxocara spp. eggs in soil and sand samples

BackgroundToxocara canis and Toxocara cati are globally distributed roundworms and causative agents of human toxocariasis, via ingestion of Toxocara eggs. Control of Toxocara infections is constrained by a lack of sensitive methods for screening of animal faeces and environmental samples potentially contaminated by Toxocara eggs. In this work, a pre-analytical method for efficient extraction of DNA from Toxocara eggs in environmental samples was set up using our previously validated T. canis- and T. cati-specific quantitative real-time polymerase chain reaction (qPCR). For this purpose, the influence of different methods for egg lysis, DNA extraction and purification for removal of PCR inhibitors were assessed on environmental samples.MethodsTo select the best egg disruption method, six protocols were compared on pure T. canis egg suspensions, including enzymatic lysis and thermal or mechanical disruption. Based on the selected best method, an analytical workflow was set up to compare two DNA extraction methods (FastDNA™ SPIN Kit for Soil versus DNeasy® PowerMax® Soil Kit) with an optional dilution and/or clean-up (Agencourt® AMPure®) step. This workflow was evaluated on 10-g soil and 10-g sand samples spiked with egg suspensions of T. canis (tenfold dilutions of 104 eggs in triplicate). The capacity of the different methods, used alone or in combination, to increase the ratio of positive tests was assessed. The resulting optimal workflow for processing spiked soil samples was then tested on environmental soil samples and compared with the conventional flotation-centrifugation and microscopic examination of Toxocara eggs.ResultsThe most effective DNA extraction method for Toxocara eggs in soil samples consisted in the combination of mechanical lysis of eggs using beads, followed by DNA extraction with the DNeasy® PowerMax® Soil Kit, and completed with an additional DNA clean-up step with AMPure® beads and a sample DNA dilution (1:10). This workflow exhibited a limit of detection of 4 and 46 T.canis eggs in 10-g sand and 10-g soil samples, respectively.ConclusionsThe pre-analytical flow process developed here combined with qPCR represents an improved, potentially automatable, and cost-effective method for the surveillance of Toxocara contamination in the environment.Graphical

Development of a Rapid and Low-Cost Method for the Extraction of Dermatophyte DNA.

Background:Polymerase chain reaction (PCR) is the most optimized method for the rapid detection and analysis of any environmental or clinically significant organism. While PCR amplification directly from samples has been shown effective for several bacteria and viruses, for filamentous fungus and yeast, extraction of genomic DNA is a must. The extraction of DNA from fungal cultures is often reported using user-friendly commercially available kits, which are designed to decrease the time, extensive manual work in extraction procedures but are often expensive. Dermatophytes pose an added drawback to efficient DNA extraction due to their poor recovery on culture media and slow growth rate.Aims and Objectives:In the present study, we developed and validated a method for effective genomic DNA extraction from dermatophytes.Materials and Methods:DNA yield from standard dermatophytes extracted from spore suspensions and mycelia mat by commercially available kits was compared. A modified method using lyticase buffer and phenol-chloroform extraction was developed. The yield obtained was compared with the existing methods (kit-based method and cetyl trimethyl ammonium bromide method). The yield and quality of the total genomic DNA were estimated spectrophotometrically and by successful PCR amplification of the ITS region. The results were validated using 21 clinical isolates from recalcitrant dermatophytosis.Results:Minimal fungal DNA was obtained from the spores compared to that obtained from mycelial mat. Commercially available kits yielded lower amounts of DNA compared to the CATB method. The modified method developed in this study yielded better quality and quantity of DNA.Conclusion:Of the three extraction methods evaluated, the developed method gave significantly higher total genomic DNA yield and better purity than the reference methods. In addition, the turnaround time for DNA extraction was reduced to half based on modifications in culture conditions.

Using acetone for rapid PCR-amplifiable DNA extraction from recalcitrant woody plant taxa.

PremiseQuick and effective DNA extraction from plants for subsequent PCR amplification is sometimes challenging when working across diverse plant taxa that may contain a variety of inhibitory compounds. Time‐consuming methods may be needed to overcome these inhibitory effects as well as the effects of various preservation and collection methods to extract DNA from leaf samples. Our objective was to develop a rapid DNA extraction protocol that could be used with diverse plant taxa to produce high‐quality DNA suitable for downstream PCR applications.Methods and ResultsWe tested the efficacy of acetone in extracting DNA from fresh, frozen, oven‐dried, acetone‐fixed, and herbarium leaf material of 22 species from 16 woody and herbaceous plant families. An improved simplified DNA extraction protocol was developed using acetone‐fixed leaf material. The addition of 1% sodium dodecyl sulfate solution resulted in the optimal extraction from all tissue samples. The DNA resulting from the extraction protocol was readily amplified using real‐time PCR assays.ConclusionsThe protocol described here resulted in the extraction of DNA from recalcitrant plant species that was of sufficient quality and quantity for PCR amplification, as indicated by the low threshold cycle values from real‐time assays. This method is simple, fast, and cost‐effective, and is a reliable tool for extracting high‐quality DNA from plant material containing PCR inhibitors.

Development of an Efficient Method to Extract DNA from Refined Soybean Oil

Recently, soybean oil has become the most consumed genetically modified (GM) vegetable oil globally. To monitor products derived from GM sources, DNA-based analysis methods are being widely adopted, particularly to control the authenticity of several food products including oil. Considering that DNA isolation from foodstuffs is the first step in the detection of GM organisms, we compared five different methods to extract high quality DNA from refined soybean oil. We used ultraviolet spectrophotometry, nested polymerase chain reaction (PCR), and real-time PCR to evaluate DNA yield and purity, and the sensitivity of the optimized method was evaluated. According to our results, the most effective DNA extraction method was the cetyltrimethylammonium bromide–NucleoSpin Food Kit method. Using this method, 21–225 DNA copies were isolated from 1 mL soybean or blended oils, and the minimum ratio of soybean oil that could be detected from a mixture of soybean and peanut oil was 5%. To summarize, we report a novel method that combines pre-concentrating oils and commercial kits. This optimized DNA extraction method advances the traceability of soybean oils for authenticity issues and transgenic detection.

An Effective and Viable DNA Extraction Protocol for FFPE Tissues and its Effect on Downstream Molecular Application

Archival Formalin-Fixed Paraffin-Embedded (FFPE) tissues are a readily available source of DNA for retrospective studies including genetic predisposition. FFPE tissues present significant challenges for extraction of pure genomic DNA in adequate amounts due to the harsh fixation conditions and long-term storage associated with samples. The aim of the study is to propose a simple, in-expensive, viable protocol for the extraction of DNA from FFPE tissues and to optimize the same for the detection of 3’UTR miRNA binding site polymorphism in ADAMTS1 gene in colorectal cancer tissues and normal colonic mucosa archival tissues using Sanger sequencing and TaqMan SNP genotyping assay. Five different DNA extraction methods were compared and analyzed. NanoDrop quantification demonstrated that out of the various methods tested, highest quantity and pure DNA was obtained from Modified non heating extraction. When we further applied this method for colorectal cancer tissues and normal colonic mucosa archival tissues the NanoDrop readout indicated that the DNA was intact and pure. Intra samples comparison over a time period of 6 months revealed that the DNA was intact. The samples isolated with Modified non heating extraction were found suitable for downstream processes such as Sanger sequencing and TaqMan SNP genotyping assay. This cost effective and non-arduous protocol can successfully extract even minute quantities of DNA from FFPE tissues and facilitate the downstream molecular analysis of a large number of archival specimens for retrospective studies.

Comparison of two DNA extraction methods: PrepFiler® BTA and modified PCI-silica based for DNA analysis from bone

Bones are important evidence often found in crime scene associated with mass grave disaster and missing persons. Substantial fragmentation and intermixing of bones limit the use of morphological observation. Human individualization using DNA analysis can complement but the challenges are the PCR inhibitors contained and low amount of DNA in degraded bone. The selection of effective DNA extraction method that minimizes PCR inhibitors and maximizes DNA recovery thus is crucial. This study aimed to compare the efficiency of the currently modified PCI-silica and the latest forensic commercial kit developed for challenging bone sample (PrepFiler® BTA). The results showed that PrepFiler® BTA provided significantly higher median DNA concentration (p < 0.05) and 95.3% probability of obtaining more alleles compared to modified PCI-silica based method. The efficiency of PCR inhibitors removal showed that both extraction methods were comparable at minimizing PCR inhibitors. Our study could serve as another guideline for selection of most suitable method for DNA analysis from bone.