Use of FFPE-derived DNA in next-generation sequencing: DNA extraction methods. (2023)

author:McDonough SJ, Bhagwate A, Sun Z, Wang C, Zschunke M, Gorman JA, Kopp KJ, Cunningham JM

Publications: plus one, 2019,rollo. 14, side e0211400

PubMed number:30973937PubMed Review papers?No

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the purpose of the thesis

This article examines the differences between DNA extraction methods in terms of DNA yield, percentage of double-stranded DNA (dsDNA), and fragment size, and compares the performance of extraction and library preparation methods in sequencing measurements. Next Generation (NGS) and file usage. Formalin-fixed paraffin Effect of concordance of genotypes in embedded samples (FFPE). Differences between tissue types were also investigated.

the conclusion of the article

DNA yield was highest in tonsil samples and lowest in brainstem and breast samples, but there were no consistent differences in yield between extraction methods. The percentage of double-stranded DNA was higher with the QIAamp DNA Mini Kit, the GeneRead DNA FFPE Kit (automated or manual), the QIAsymphony DSP DNA Mini Kit, and the KingFisher Duo MagMAX FFPE DNA Isolation Kit. Get the most consistent intact DNA (highest DQN and best multiplex PCR) with the QIAamp DNA Mini Kit, QIAcube GeneRead DNA FFPE Kit, and KingFisher Duo MagMAX FFPE DNA Isolation Kit.

(Video) Webinar - Extraction Strategies for comprehensive DNA and RNA analysis from the same FFPE sample 🧬

As expected, the most degenerate sample (cerebellum) had lower average coverage and higher replication rates than the other tissue types. Using Ultra II instead of ThruPLEX for library preparation resulted in higher average coverage and lower replication rates, but the difference in extraction method was not significant. Genotype agreement between extraction sets was >95% for all extraction set combinations and both library preparation methods.

With the specific NGS panel, the highest average coverage was recorded in samples extracted with the KingFisher Duo MagMAX FFPE DNA Isolation Kit. The average agreement between genotype call extraction methods using specific panels was 99.59%. The C>T/G>A percentages were lower when extracted with the QIAcube GeneRead DNA FFPE Kit or the KingFisher Duo MagMAX FFPE DNA Isolation Kit, but the difference was small.


  1. learning purpose

    This study examined differences between DNA extraction methods in terms of DNA yield, dsDNA percentage, and fragment size, and compared extraction and library preparation methods in terms of NGS metrics and genotype agreement. The effect of tissue type was also investigated. Nine 10 µm sections were dissected from each of 12 archived FFPE blocks, including normal and pathological breast, colon, lung and pancreas specimens; two normal tonsil tissues; brainstem samples; and cerebellar samples. Samples were fixed for more than 6 h before paraffin embedding, but other details of sample collection and FFPE processing were not specified. Sections were placed on glass slides and scraped into microcentrifuge tubes. Deparaffinize sections with QIagen Deparaffinization Solution, xylene (for QIAamp extraction) or KAPA Express Extract Reagent (for KAPA Rapid Extraction). Use manual kits (KAPA Express Extraction Kit, Promega Reliaprep FFPE gDNA Miniprep System, QIAamp FFPE Tissue Kit with Overnight Assay Extension, or QIAGEN GeneRead DNA FFPE Kit) or automated kits (QIAGEN QIAsymphony DNA Mini Kit, QIAGEN GeneRead DNA) DNA Extract FFPE Kit, Maxwell RSC DNA FFPE Kit, PerkinElmer Chemagic FFPE DNA Kit, or Applied Biosystems Duo Mag-MAX FFPE DNA Isolation Kit). DNA was quantified using the Qubit dsDNA BR assay and a spectrophotometer. For samples measuring less than 2.0 ng, the concentration was verified using the Qubit dsDNA HS assay. Fragment lengths were determined using the Analytical Fragment Analyzer High Sensitivity Large Fragment Analysis Kit and multiplex PCR. Libraries were constructed with NEBNext Ultra II DNA Library Prep and ThruPLEX DNA-seq Kit from DNA extracted with QIAcube GeneRead DNA FFPE Kit (manual and automated), QIAamp DNA Mini Kit, or KingFisher Duo MagMAX FFPE DNA Isolation Kit and sequenced on Illumina HiSeq 4000. The QIAGEN QIAseq Comprehensive Targeted Human Cancer Panel and Human Breast Cancer Panel also sequenced these samples with the HiSeq 4000. Statistical analyzes were performed, but comparisons were not always clear cut.

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    Summary of Results:

    DNA yield was highest in tonsil samples and lowest in brainstem and breast samples, but extraction method did not uniformly affect yield. As expected, the tumors produced more DNA than the corresponding normal tissues. The percentage of double-stranded DNA was higher with the QIAamp DNA Mini Kit, the GeneRead DNA FFPE Kit (automated or manual), the QIAsymphony DSP DNA Mini Kit, and the KingFisher Duo MagMAX FFPE DNA Isolation Kit. Absorbance indices of 260 to 280 were greater than 1.8 for all samples except samples extracted with the KAPA Express Extract Kit and the Reliaprep FFPE gDNA Miniprep System. Fragment length analysis showed the most severe degradation in cerebellar samples and using the KAPA Express Extract Kit, Reliaprep FFPE gDNA Miniprep System, Maxwell RSC DNA FFPE Kit, or Chemagic MSM1 FFPE DNA Kit. Highly variable results were observed with the QIAsymphony DSP DNA Mini Kit. The most consistent intact DNA (highest DQN) was obtained using the QIAamp DNA Mini, QIAcube GeneRead DNA FFPE, and KingFisher Duo MagMAX FFPE DNA Isolation Kits. Multiplex PCR confirmed that samples extracted with KAPA Express Extract, Chemagic MSM1 FFPE DNA, and QIAsymphony DSP DNA Mini kits had lower integrity, while samples extracted with QIAcube GeneRead DNA FFPE Kit or KingFisher Duo MagMAX FFPE DNA Isolation Kit they had more time The amplification effect of the fragment is better

    As expected, the most degenerate sample (cerebellum) had lower average coverage and higher replication rates than the other tissue types. Using Ultra II instead of ThruPLEX for library preparation resulted in higher mean coverage (mean 25.88X vs. 21.5X) and lower doubling rates (11.9% vs. 26.21). %). There were no significant differences in mean coverage or replicates between extraction methods, and genotype agreement between extraction sets was >95% for all extraction set combinations and for both library preparation methods.

    Using the QIAseq Human Comprehensive Cancer panel, samples extracted with the KingFisher Duo MagMAX FFPE DNA Isolation Kit (820.91X) were found to have higher Average Molecular Label Coverage) or the QIAamp DNA Mini Kit (599.68X). Likewise, for samples extracted with the KingFisher Duo MagMAX FFPE DNA Isolation Kit (1609.23X), coverage was higher with the breast cancer panel than with the QIAamp DNA Mini Kit (1089.31X) or the QIAcube GeneRead DNA FFPE Kit manual. or automated (749.12) X), X) and 731.92X respectively). Importantly, the average concordance between genotype calls between extraction methods was 99.59%. Finally, the C>T/G>A percentages were lower when extracted with the QIAcube GeneRead DNA FFPE Kit (36.64%) or the KingFisher Duo MagMAX FFPE DNA Isolation Kit (36.59%), while extracted with the QIAamp DNA, C>T/G>A has the highest percentage of minisets (38.06%).

    (Video) Techniques and Tips: Learn Sample Preparation for Next Generation Sequencing

    biological sample
    • tissue - brain
    • tissue - breasts
    • Tissue - tonsils
    • tissue - lung
    • Tissue - Pancreas
    • Tissue - Colorectal
    types of preservatives
    • Formalin
    to diagnose:
    • normal
    • tumor - cancer
    analyteTechnological platform
    decalcified nucleosaccharide nucleic acid fluorescent decalcified nucleosaccharide nucleic acid next generation sequencing decalcified nucleosaccharide nucleic acid Polymerase chain reaction decalcified nucleosaccharide nucleic acid spectrophotometry
    Factors before analysis:
    classificationpreanalytical factorsvalor
    Extraction and purification of analytes Separation of analysis KAPA quick release kit
    Promega Reliaprep FFPE gDNA minipreparation system
    QIAamp FFPE Tissue Kit (lysis time extended to overnight)
    Folleto del kit QIAGEN GeneRead DNA FFPE
    QIAGEN GeneRead DNA FFPE Automated Kit
    QIAsymphony DNA mini kit from QIAGEN
    Kit Maxwell RSC ADN FFPE
    PerkinElmer Chemical FFPE DNA Kit
    Applied Biosystem Duo Mag-MAX FFPE DNA Isolation Kit
    collection of biological samples biological sample location brainstem
    the cerebellum
    the pancreas
    Next Generation Specific Sequence Technological platform Comprehensive QIAseq-led human cancer panel
    Human Breast Cancer Kit
    NEBNext Ultra II DNA Library Preparation
    ThruPLEX DNA Sequencing Kit

    see more

    (Video) Optimizing FFPE Extraction for Methylation Array and Sequencing Assays - AMP 2022 Workshop - FULL

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How do you extract DNA from FFPE tissue? ›

To extract DNA from paraffin-embedded samples, tissue cores or microdissected tissue are subjected to xylene treatment, which dissolves the paraffin from the tissue, and then rehydrated using a series of ethanol washes.

What is DNA extraction for NGS sequencing? ›

Next-generation sequencing: DNA extraction protocol

Extraction entails breaking down the extracellular matrix and opening the cell membranes using enzymes, solvents, or surfactants. The DNA in the resulting mixture must then be isolated.

What does FFPE stand for? ›

Formalin-Fixed Paraffin-Embedded (FFPE) tissue specimens have been a staple of research and therapeutic applications for decades. FFPE is a form of preservation and preparation for biopsy specimens that aids in examination, experimental research, and diagnostic/drug development.

How much tissue is required for next-generation sequencing? ›

Regardless of the device used, a tissue surface area of ≥ 1 mm2 is adequate for samples to be tested with NGS. Keywords: Oncomine Dx Target Test; bronchoscopy; next-generation sequencing; non-small-cell lung cancer.

What are the steps of tissue processing of FFPE? ›

Steps for FFPE Tissue Preparation
  • Fresh Tissue. First, fresh tissue must be obtained. ...
  • Liquid Fixing Agent. ...
  • Dehydration by Ethanol. ...
  • Clearing Agent. ...
  • Paraffin Wax.

How do you extract DNA from a tissue sample? ›

Preparing the Tissue Lysate
  1. Place the tissue sample into a sterile microcentrifuge tube.
  2. Add 1 ml of Lysis Mix (see above) to the tube. ...
  3. Vortex for 10-15 seconds to mix.
  4. Incubate the sample overnight at 55° C until lysis is complete. ...
  5. Add 5 µl of RNase A to the lysate. ...
  6. Incubate at room temperature for 5 minutes.

What is the difference between NGS and DNA sequencing? ›

The critical difference between Sanger sequencing and NGS is sequencing volume. While the Sanger method only sequences a single DNA fragment at a time, NGS is massively parallel, sequencing millions of fragments simultaneously per run. This process translates into sequencing hundreds to thousands of genes at one time.

What are the different types of NGS DNA sequencing? ›

There are three main types of DNA sequencing done by NGS: Whole Genome Sequencing, Whole Exome, and Targeted Panels.

What is the function of FFPE? ›

Formalin-Fixed Paraffin-Embedded (FFPE) scrolls can be used to extract DNA and can be used in whole-genome sequencing and other genomic applications for pipeline validation and assay development.

What is FFPE DNA? ›

Formalin-fixedparaffin-embedded (FFPE) tissues are a very common source of archived tissue samples that have been used by pathologists for decades. It allows for a long-term, room temperature storage solution for tissue samples, making it very convenient.

What does FFPE mean in pathology? ›

Abstract. Paraffin embedding is a standard technique used in clinical and research laboratories to create a formalin-fixed, paraffin-embedded (FFPE) block of tissue. Formalin-fixed tissue undergoes tissue processing and then is embedded in paraffin (wax) to create a FFPE block or paraffin block.

How is next-generation sequencing NGS most often used? ›

Unlike DNA sequencing, this is called RNA sequencing. Specially designed mRNA sequencing is also often used to detect fusion genes. The most commonly used NGS assay for cancer patients is targeted panel sequencing which usually interrogates dozens or hundreds of targeted genes.

How much does next-generation sequencing NGS cost today? ›

The cost of a human genome sequence decreased from an estimated $1 million in 2007, to $1000 in 2014, and today it is approximately $600.

Why is next-generation sequencing cheaper? ›

Since the completion of the Human Genome Project, the cost of next-generation sequencing (NGS) has decreased at a dramatic rate, outpacing Moore's Law. Through continuous innovation, Illumina has helped reduce the cost of NGS, enabling the $1000 human genome.

What temperature should FFPE be stored at? ›

Question: How should I store my FFPE tissue blocks? Answer: FFPE blocks are typically stored at room temperature but storage at 4°C and protected from light is optimal for 10x Assays as this will slow the degradation of RNA.

What are the three main methods of tissue processing? ›

Tissue processing is the technique by which fixed tissues are made suitable for embedding within a supportive medium such as paraffin, and consists of three sequential steps: dehydration, clearing, and infiltration.

How long can tissue be stored in formalin? ›

All tissue specimens should be fixed in 10% neutral buffered formalin for a minimum of 3 days (72 hours) and up to 2 weeks (14 days) for biopsy or autopsy tissue, or up to 4 weeks (28 days) for brain autopsy tissue.

How do you know if DNA extraction is successful? ›

The absorbance quotient (OD260/OD280) provides an estimate of DNA purity. An absorbance quotient value of 1.8 < ratio (R) < 2.0 was considered to be good, purified DNA. A ratio of <1.8 is indicative of protein contamination, where as a ratio of >2.0 indicates RNA contamination.

What is the best way to extract DNA? ›

Manual methods as well as commercially available kits are used for DNA extraction. Various tissues including blood, body fluids, direct Fine needle aspiration cytology (FNAC) aspirate, formalin-fixed paraffin-embedded tissues, frozen tissue section, etc., can be used for DNA extraction.

What are the three steps of DNA extraction? ›

DNA extraction is the process where DNA is separated from proteins, membranes, and other cellular material (Butler, 2012). According to Rice (2018), the method involves three necessary steps, namely, lysed, precipitation, and purification.

What are the disadvantages of NGS sequencing? ›

The limitations of NGS in clinical microbiology include unclear antibiotic resistance, inability to detect which pathogen is causing disease, costs, and unknown clinical utility.

How accurate is NGS sequencing? ›

Among NGS platforms, the accuracy of the original base data obtained by SOLiD platform is greater than 99.94%, though accuracy can reach 99.999% with the sequencing depth of 15×, which is the highest accuracy in NGS platforms (Ronchi et al., 2012).

What is the most accurate sequencing method? ›

With over 99% accuracy, the Sanger sequencing method remains the “gold standard” for basic and clinical research applications. In fact, most clinical laboratories rely on Sanger sequencing to validate gene variants (e.g., single-nucleotide variants and insertion/deletions) identified first through NGS.

What is the most popular DNA sequencing method? ›

Due to its sensitivity and relative simplicity in terms of both workflow and technique, Sanger sequencing remains the gold standard in sequencing technology today and is used in a variety of applications from targeted seqencing to confirming variants identified using orthogonal methods.

What is the most common NGS? ›

Illumina, Nanopore, and PacBio are among the most commonly used high-throughput sequencing platforms (Ye et al., 2015) . ...

Why do we need next-generation sequencing? ›

Key Benefits of NGS

Using capillary electrophoresis-based Sanger sequencing, the Human Genome Project took over 10 years and cost nearly $3 billion. Next-generation sequencing, in contrast, makes large-scale whole-genome sequencing (WGS) accessible and practical for the average researcher.

What are the most important steps in DNA extraction protocol? ›

There are 3 basic steps involved in DNA extraction, that is, lysis, precipitation and purification. In lysis, the nucleus and the cell are broken open, thus releasing DNA. This process involves mechanical disruption and uses enzymes and detergents like Proteinase K to dissolve the cellular proteins and free DNA.

Why is DNA extraction important in the process? ›

The ability to extract DNA is of primary importance to studying the genetic causes of disease and for the development of diagnostics and drugs. It is also essential for carrying out forensic science, sequencing genomes, detecting bacteria and viruses in the environment and for determining paternity.

What precautions should be taken during DNA extraction? ›

Wear personal protective equipment, such as a lab coat, gloves and eye protection. Do not get into eyes, on skin or clothing. Do not breathe in vapours or dust from dried-up solution. Do not ingest.

How can I improve my FFPE DNA quality? ›

Therefore, formalin fixation is a major cause of reduced quality of DNA from FFPE tissues, and heat treatment after lysis can dramatically improve the quality.

How long can FFPE sections be stored? ›

Schlumpberger: Generally, we recommend storing FFPE sections or blocks cold at 4° or -20°. At -20°, blocks can be used after ten years. Eventually, there will be a decline in integrity.

Why is tissue fixation needed? ›

Fixation is the foundation step behind the study of pathology and essentially exists to prevent the autolysis and degradation of the tissue and tissue components such that they can be observed both anatomically and microscopically following sectioning.

What is the yield of DNA FFPE extraction? ›

Based on the results from the assessment of nucleic acid extraction kits, nucleic acids were extracted using the QIAamp® DNA FFPE Tissue kit and RNeasy® FFPE kit. DNA was successfully extracted from all 84 biopsies with a mean yield of 3.7 ng/μl and a purity of 1.7.

What are other methods suitable for isolation of DNA from FFPE or FF tissues? ›

A standard method to isolate DNA from tissues is by tissue digestion with proteinase K (PK), followed by heating to 100°C to inactivate the enzyme. After treatment with RNAse, genomic DNA can then be recovered by salt precipitation or with a resin column for downstream applications (Green and Sambrook, 2012‬).

What is the effect of formalin on DNA? ›

For molecular analysis, the duration of fixation is the major factor affecting the preservation of DNA in formalin fixed tissue. Unbuffered formalin oxidises to formic acid and an acidic environment causes degradation of nucleic acids because the β glycosidic bonds in the purine bases are hydrolysed at pH 4.

What is the purpose of tissue processing in histopathology? ›

The tissue processor finds applications in histopathology laboratories to automatically prepare tissue samples for laboratory testing, by fixing, dehydrating, clearing, and infiltrating them with paraffin.

What chemicals are used to fix FFPE tissue? ›

Solution. The fixative that is used by most laboratories is neutral-buffered, 10% formalin. Formalin is a liquid that is 37-40% formaldehyde and 10% methanol (by weight) in water; thus a 10% formalin solution contains about 3.7% – 4% formaldehyde and 1% methanol (Hewitt, et al., 2008; Kiernan, 2000).

What percentage of tumors are in FFPE? ›

The average tumor cell content was 70% in fresh frozen and 68% in FFPE samples.

What is the biggest challenge for next-generation sequencing NGS? ›

In next-generation sequencing workflows, samples of low or variable quality can corrupt downstream processes such as library preparation and ultimately confound analysis. Samples should be assessed for crosslinks, breaks, the accumulation of single-stranded DNA, and other forms of damage.

What is the effectiveness of NGS? ›

NGS is a cost-effective strategy compared with single-gene testing (SgT) in the molecular diagnosis of patients with metastatic NSCLC. More targetable genomic alterations could be detected by NGS, and therefore, more patients could potentially be treated with targeted therapies or enrolled in specific clinical trials.

What is next-generation sequencing in simple words? ›

Next generation sequencing (NGS), massively parallel or deep sequencing are related terms that describe a DNA sequencing technology which has revolutionised genomic research. Using NGS an entire human genome can be sequenced within a single day.

Is NGS covered by Medicare? ›

Specifically, the Centers for Medicare & Medicaid Services (CMS) established coverage of NGS as a diagnostic laboratory test when performed in a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory, ordered by a treating physician, and specific criteria are met.

Is next-generation sequencing FDA approved? ›

FDA Approves Liquid Biopsy Next-Generation Sequencing Companion Diagnostic Test. On August 26, 2020, the Food and Drug Administration approved the liquid biopsy next-generation sequencing-based FoundationOne Liquid CDx test (Foundation Medicine, Inc.)

How long does NGS sequencing take? ›

The testing typically takes 2 to 3 weeks for results, while individual tests often take a week or less. The cost of testing (typically several thousand dollars) is more than the cost of testing one or a few markers, if no more are needed.

What are the two types of next-generation sequencing? ›

There are three main types of DNA sequencing done by NGS: Whole Genome Sequencing, Whole Exome, and Targeted Panels. Choosing between the type of NGS that may be required for research or clinical diagnosis depends on what you are looking for.

What is the latest technology for DNA sequencing? ›

Researchers have created a new DNA sequencing technique called Chem-map, which enables researchers to perform in situ mapping of small molecule-genome interactions with unparalleled precision.

What is the error rate of next-generation sequencing? ›

Conventional intrinsic NGS error rates range between 0.1 and 1% (Phred quality score of 20–30) (1, 11) depending on the sequencing platform, the GC content of the target regions (12), and the fragment length, as shown in Illumina paired-end sequencing (13).

How is DNA released from silica membrane? ›

Often, DNA elution is facilitated by high temperature, high pH, and low ionic strength conditions. Higher pH conditions likely facilitate DNA elution by increasing the negative charge density on the silica surface, resulting in greater electrostatic repulsion between the DNA and silica surface.

Can DNA be extracted from dead tissue? ›

With deceased and decayed bodies, personal identification is performed using hard tissue DNA, commonly extracted from bone. The quantity and quality of DNA used in the polymerase chain reaction (PCR) amplification step is critical for a successful outcome.

Can DNA be extracted from dried material? ›

The quality of the DNA extracted from the samples was measured. Results No difference was observed between fresh and stored blood samples. Conclusions The quality of DNA extracted from liquid or dried blood is not adversely affected by storage at 4° C for up to 24 h.

How do you extract DNA step by step? ›

There are 3 basic steps involved in DNA extraction, that is, lysis, precipitation and purification. In lysis, the nucleus and the cell are broken open, thus releasing DNA. This process involves mechanical disruption and uses enzymes and detergents like Proteinase K to dissolve the cellular proteins and free DNA.

How much material is needed to extract DNA from your sample? ›

For DNA sequencing, the amount is dependent upon the type of the sample you have. Soil and tissue samples require at least 0.25g. We recommend sending a few grams extra especially if multiple extraction replicates per sample are requested.

What is the purpose of DNA extraction? ›

The purpose of genomic DNA extraction is to separate this genetic material from the rest of the cell (proteins, RNA, cell membrane, etc.). Once purified, scientists can study individual genes, sequence the entire genome, modify sections of DNA, and more.

How long does it take to analyze DNA after extraction? ›

You can expect your results within eight weeks of the date the laboratory receives your sample.

How long does DNA last on a tissue? ›

About a month to a million years, theoretically. The decay rate of DNA depends on the conditions of its storage and packaging. Above all, it depends on whether the DNA is exposed to heat, water, sunlight, and oxygen.

How do you preserve tissue for DNA extraction? ›

We recommend freeze drying of DNA samples, especially when they have to be shipped for longer distances. No special packaging or declaration is needed for freeze dried samples, and the risk of thawing is excluded.

Does it matter what type of tissue DNA is extracted from? ›

No, it does not matter what tissue is used in an organism, the DNA sequence is the same in all cells of an organism. One must keep in mind that in blood, red blood cells do not have a nucleus, therefore, no DNA. When a DNA extraction process is done on blood, the DNA actually comes from the white blood cells.

What is the best tissue for DNA extraction? ›

It is suggested that the brain tissue preserved at −80°C is the best among soft issues for DNA extraction.

How can we prevent DNA degradation during extraction? ›

As any enzyme activity is sensitive to temperature, the degradation process is reduced at lower temperatures. Thus, keeping samples cold will slow down the enzymatic degradation of DNA.

Can I freeze whole blood for DNA extraction? ›

Cryopreservation of whole blood is useful for DNA collection, and clinical and basic research. Blood samples in ethylenediaminetetraacetic acid disodium salt (EDTA) tubes stored at - 80 °C are suitable for DNA extraction, but not for high-quality RNA extraction.

What are 5 ways DNA can be extracted? ›

Basic Isolation Procedure
  • Physical Methods. Physical methods typically involve some type of sample grinding or crushing to disrupt the cell walls or tough tissue. ...
  • Chemical Methods. ...
  • Enzymatic Methods. ...
  • Solution-Based Chemistry. ...
  • Silica-Binding Chemistry.

What is the easiest way to collect DNA? ›

For DNA testing the most popular and reliable way to collect samples is the oral buccal swab method. A buccal swab closely resembles a one ended Q-Tip in appearance. Using swabs as a collection method is quick and painless and is the recommended way to collect DNA samples for testing.

Why is salt added in DNA extraction? ›

By adding salt, we help neutralize the DNA charge and make the molecule less hydrophilic, meaning it becomes less soluble in water. The salt also helps to remove proteins that are bound to the DNA and to keep the proteins dissolved in the water.


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