EzWay Direct PCR Buffer

EzWay Direct PCR Buffer (5x)

Catalog Number	K0568001
Size	500ul
Type	PCR Kit
Datasheet

Description

EzWay™ Direct PCR Buffer enables the direct amplification of DNA fragments from a variety of samples including whole blood, blood collected on card, saliva, bacteria, mouse tail, tissue, cultured cells or plant eliminating the need for DNA purification, just by adding to the PCR mixture in place of general 10X PCR Buffer.

EzWay™ Direct PCR Buffer is compatible with most commercially available any DNA polymerases including Taq, Pfu, Pwo, Tth and even with any type of Hot Start Taq DNA polymerase. A one-step reaction using this EzWay™ Direct PCR Buffer enables easy PCR and minimizes the sample cross-contamination when dealing with multiple tissue or samples by eliminating tedious genomic DNA preparation.

Features

No need to purify DNA prior to PCR
Minimizes the risk of DNA contamination during reagent handling
Prevents the loss of trace samples in the DNA purification process
Amplification of DNA directly from samples containing : Whole Blood, Blood Stains, Blood Cards, Buccal Swab, Saliva, Hair root, Sperm, Body Fluid, Cultured Cells, Bacteria, Viruses, and Tissues (fresh, frozen or paraffinized), Plant etc.
Compatible with various sources of thermostable DNA polymerases

Application

Viral infection / Molecular diagnostic test
Forensic DNA Analysis
Single cell diagnostic
Blood banking
Identity testing
Multiplex PCR / SNP detection / PCR-RFLP
Sequencing / Cloning
Laboratory Automatic PCR

Storage

6 months at 4°C, 1 year at -20°C

Packing

Component	Cat.No	Volume	Comment
5X EzWay™ Direct PCR Buffer*	K0568001 or K0568011	500 ul	Final 1.5mM MgCl₂

Note:

Most of commercially available Taq Polymerases can be used with K0568001 or K0568002 for Direct PCR application, but in case of AmpliTaq Gold (from Applied Biosystems), K0568011 or K0568012 is highly recommended to be used for the best performance.
Direct Lysis Buffer I/II, Direct Elution Buffer, or Magic Buffer are not included in this product, so they must be purchased or prepared separately for the Lysis Method. EzWay™ Direct PCR Buffer Set (K0568002 or K0568022) is recommended to use for application to the samples requiring the Lysis or Elution process.

Fig.1 Genomic DNA PCR vs. Blood Direct PCR of different sizes of p53 targets

PCR amplifications were performed using 20 ng human genomic DNA or 1 ul Heparin-treated whole blood in a 50ul PCR reaction volume. EzWay™ Direct PCR Buffer can amplify directly up to 2kb amplicon and produce more specific band patterns than DNA PCR. It means that direct PCR amplification has inherent hot start function even without the help of hot start enzyme because the cells are disrupted in initial heating stage, then the exposed DNA templates meet primers instantly at higher temperature than Tm.

Fig.2 Direct PCR of p53 from human whole blood Blood treated with three anti-coagulants

Direct amplification from anticoagulants treated human blood with EzWay™ Direct PCR Buffer (Lane2-7) and conventional PCR buffer (Lane1). The inhibitory effects caused by anti-coagulants and blood components is successfully overcome and PCR result can be achieved even in the condition of maximum 10-20%(v/v) blood volume in a total reaction volume.
Lane P: Human genomic DNA, Lane N: Negative controls

Fig.3 Direct sequencing from DNA and EDTA-blood Apolipoprotein E genotyping

Apolipoprotein E, e2/e4, is identified with DNA (A) and EDTA-blood (B) by PCR and direct sequencing using BigDye® Terminator v3.1 cycle sequencing reaction. The amplicon from blood by EzWay™ Direct PCR Buffer can be used as sequencing template. Then, the direct PCR chemistry does not interfere downstream dideoxy sequencing reaction and fluorescent detection.

Fig.4 AmpliTaq Gold specific Direct PCR Buffer Gold

Blood having 5 different genotypes (Lane 1,6,11, Lane 2,7,12, Lane 3,8,13, Lane 4,9,14 and Lane 5,10,15, respectively) was amplified by Direct PCR method. The Direct PCR Buffer Gold is specific to AmpliTaq Gold DNA Polymerase as shown at the lane 6-10. AmpliTaq Gold didn't work well when it was used with general type of Direct PCR Buffer (lane 16-20).
Lane 1-5 : Direct PCR using Direct PCR Buffer Gold (K0568011) with KOMA Taq DNA Polymerase
Lane 6-10 : Direct PCR using Direct PCR Buffer Gold (K0568011) with AmpliTaq Gold DNA Polymerase
Lane 11-15 : Direct PCR using Direct PCR Buffer (K0568001) with KOMA Taq DNA Polymerase
Lane 16-20 : Direct PCR using Direct PCR Buffer (K0568001) with AmpliTaq Gold DNA Polymerase