BEAMSBIO GS-Taq PCR Master Mix (2X)

Store at –20℃ for storage, but 4℃ for daily or weekly use
#3308 5ml	4vials of 1.25 ml/vial : 5 ml for 500 reactions of 20 µl. Supplied with 0.3 ml of loading dye(x6)
* Recipe : 80 U/ml GS-Taq DNA Polymerase, 1.6 mM dNTP mix & 2x Reaction buffer

Features

l         Taq DNA Polymerase fortified with High Fidelity and 3’à5’ proofreading activity

l         Generate TA cloning site of PCR products.

l         Reproducible, nuclease-free, minimizing pipetting error & tedious manipulation

l         Suitable to TA cloning : PCR products with 3’-dA overhangs.

Description

PCR Master Mix (2X) is a 2X concentrated solution including GS-Taq DNA Polymerase, dNTPs and the reaction buffer for PCR. The BEAMSBIO GS-Taq PCR Master Mix(2x) facilitates PCR formulation and minimizes error and inconvenience in PCR preparation, which is suitable to difficult PCR and routine PCR for general purpose, and can amplify longer DNA template(~10kb) with greater yield and fidelity than Taq DNA Polymerase, showing more than four-times the fidelity of Taq DNA Polymerase.

Applications

l         PCR above 10KB viral DNA and 5kb genomic DNA template

l         High throughput PCR and High reproducibility with 3’à5’ proofreading activity.

l         Generation of PCR products for TA cloning.

l         RT-PCR.

Quality Control

The BEAMSBIO GS-Taq PCR Master Mix(2x) is confirmed free of endodeoxyribonucleases, exodeoxyribonucleases and ribonucleases by strict quality tests, which shows satisfactory PCR results upto 10 kb with viral DNA templates and up to 5 kb with genomic DNA templates.

Mg²⁺ conc. in PCR Master Mix

Mg²⁺ conc. is  3mM in our PCR Master Mix.  To amplify cDNA, Mg²⁺ may need to be added to a final  3mM in each PCR reaction mixture.

l       보관/사용 주의사항 :
- 최초 사용 전까지는 –20℃에 냉동 보관하십시오.
- 제품을 녹인 다음에는 반드시 5회정도 튜브를 뒤집어서 잘 섞고 나서
  사용하십시오.
- 계속 사용을 위해 냉장 보관하십시오.  3개월이상 보관이 가능합니다.

BEAMSBIO GS-Taq PCR Master Mix (2X) 매뉴얼

PCR Protocol using PCR Master Mix

1. Prepare PCR tubes for the experimental samples to be amplified.
증폭할 실험시료에 적합한 PCR튜브를 준비한다.

2. Add the components in order according to following recipe :
다음 성분표를 참조하여 실험용 재료를 순서대로 넣는다.

  Distilled water,                                                      20~24㎕ (balance to total vol. 50㎕)
  PCR Master Mix,                                               25㎕ (투입 후 가볍게 섞는다)

  DNA templates(10~200ng/㎕),                     0.2~1㎕
  Primer, forward(10pmol/㎕),                             0.2~2㎕
  Primer, backward(10pmol/㎕),         0                  .2~2㎕
-----------------------------------------------------------------------------
                                                                 total  50㎕
 * Primers(template와 미리 배합 사용가능)를 마지막에 넣는 것이 좋다
 * Magic buffer if required, 2~5㎕ *(필요 시에만 소량 사용).

3. Mix gently and rapidly. Centrifuge down PCR tubes immediately after mixing.
가볍고 신속하게 섞고, 즉시 짧게 원심분리하여 반응물을 아래로 모은다.

4. Perform PCR using optimized cycling conditions.  Suggested cycling parameters for PCR using PCR Master Mix are :
다음 운전조건을 참조하여 PCR Master Mix 최적 PCR운전조건을 설정하여 실험을 수행한다.

  - Step 1 :  initial denaturation,                                     94~5℃ for 1~3 min.
  - Step 2 :  denaturation,                                    94℃ for 30 sec.
            annealing,                                    Tm-5℃ for 45 sec.
            extension,                                    72℃ for 0.5~1 min/kb
           * 25~30 cycles recommended
           * 10KB내외인 경우, 10~15 싸이클 수행후 20~30초씩 시간을
              늘리면서 15~20 싸이클 추가로 수행함(조건 최적화 필요).
  - Step 3 :  final extension,                                    72℃ for  7~20 min.

5. Cool down below 15℃ until harvest.

PCR Troubleshooting

1. PCR band가 전혀 없는 경우 :
- Primers, template 배합에 착오가 없는지 확인한다.  성분누락 여부 ?
- PCR 운전조건이 적합하였는지 확인한다.  PCR반응기 운전 실수 여부 ?
- PCR Master Mix용도에 맞지 않는 DNA template 증폭시도 여부 ?
 Long PCR Master Mix, Pfu PCR Master Mix 등 적합한 PCR System 선택

..... 이하 여백 ... 제품매뉴얼 참조 !