DESCRIPTION

KOD One PCR master Mix and KOD One PCR Master Mix -Blue- are 2 x PCR master mixes based on genetically modified KOD DNA polymerase (UKOD). KOD One series enables fast PCR, which has an extension time of 5 sec/ kb by applying UKOD and a new Elongation Accelerator. In addition, these master mixes provide greater efficiency and elongation capabilities than conventional PCR enzymes. In particular, these show greater amplification success from crude specimens. Furthermore, these master mixes can be applied to amplify from templates containing uracils (dU) or using primers containing inosines (dI) and uracils (dU).

KOD One series contains two types of anti-KOD DNA polymerase antibodies that inhibit the polymerase and 3’→5’ exonuclease activities, thus allowing for Hot Start PCR. These master mixes generate blunt-end PCR products because of 3’ → 5’ exonuclease (proof-reading) activity of KOD DNA polymerase.

FEATURES

- Fast

KOD One series can amplify the targets using the following very short conditions:
≤1 kb: 1 sec
1~ 10 kb: 5 sec/ kb
10 kb~: 10 sec/ kb
The cycling conditions can be set flexibly when various targets having different sizes are amplified.

- Easy to Use

KOD One series contains all reaction components except primers and templates and provide high reproducibility by reducing operations. In addition, KOD One PCR Master Mix -Blue- includes a loading dye (BPB) to allow direct loading onto agarose gels.

- High Fidelity

KOD One series exhibits approximately 80-fold higher fidelity than Taq DNA polymerase. These mixes can be used for various purposes where this would be an advantage (e.g., in the preparation of long target amplicons for sequencing).

- High Efficiency

KOD One series is effective for amplification from crude samples (e.g., biological samples, foodstuffs, soil extract, etc.). Various samples or lysates can be used directly as templates.

- Primers or Templates Containing Inosines (dI) or Uracils (dU) Can Be Used

KOD One series can use primers or templates containing inosines (dI) or uracils (dU), whereas conventional high-fidelity PCR enzymes cannot.

APPLICATIONS

• Direct PCR
• Colony PCR
• Amplification of NGS libraries
• Site direct gene mutation

SOURCE

• E. coli strain carrying the cloned UKOD DNA polymerase gene

COMPONENTS

KOD One series include the following components for 200 reactions, 50 μl total reaction volume.

<KMM-101>
KOD One PCR master Mix 1 ml x 5

<KMM-201>
KOD One PCR master Mix -Blue- 1 ml x 5

Note
The reagents can be stored at 4°C for a month. For longer storage, the reagents should be kept at -20°C.

APPLICATION DATA

Example 1.Fast PCR
Various targets were amplified with KOD One PCR Master Mix and KOD One PCR Master Mix -Blue- using the ultra fast cycling conditions. KOD One series successfully amplified all targets.

Example 2.Amplification from cDNA
Inhibitory effect of RNA in cDNA was compared using various PCR enzymes. KOD One Master Mix was not susceptible to RNA inhibition, and it was able to amplify targets under high concentrations of cDNA.

Example 3.PCR error ratio
The error ratio of various PCR enzymes were compared by determining the sequences of the amplicons from human β-globin gene. The amplicons were cloned into the vector using TArget cloneTM -Plus- (Code No. TAK-201) and the sequences were determined.
KOD One PCR Master Mix and KOD One PCR Master Mix -Blue- showed excellent fidelity and the mutation frequency were approximately 80 times lower than Taq DNA polymerase.

Example 4.Amplification using degenerate primers, containing inosine(I)
The 2.8 kb fragments were amplified using degenerate primers containing inosine(I). KOD One PCR Master Mix was able to amplified, whereas conventional high-fidelity PCR enzymes cannot.

Example 5.Amplification from crude samples
Amplification from whole blood and that from mouse lysate were compared. KOD One PCR Master Mix amplified the targets efficiently.