CycleScript Reverse Transcriptase

CycleScriptTM Reverse Transcriptase는 고순도로 정제된 역전사 효소에 안정화 물질을 첨가하여, 55°C 에서도 활성이 유지되어 고온에서도 cDNA를 합성할 수 있는 제품입니다. 본 제품은 기존의 42°C 단일 온도 역전사 반응 (Fixed Temperature Reverse Transcription, FTRT) 뿐만 아니라, PCR 반응과 같이 2-3 단계의 순차적인 온도 변화를 통한 순환적 역전사 반응 (Cyclic Reverse Transcription [CRT])을 수행할 수 있는 제품입니다.

₩84,000
카탈로그 번호
E-3131-CFG

Features and Benefits

Flexible Reaction Conditions:
순환 온도 역전사 반응은 저온 (15~40℃)에서 primer annealing이 진행되고 고온 (50~55℃)에서 template RNA의 2차 구조 형성을 풀어주는 2~3개 step을 반복적으로 적용하여 기존의 42℃ 역전사 반응보다 높은 효율로 cDNA를 합성할 수 있습니다. 순환 온도 역전사 반응과 더불어 22~55°C 내의 단일 온도에서 역전사 반응도 가능합니다. 

figure1 figure1

High Stability:
안정화 물질이 포함된 건조 제품으로 RTase의 열안정성 증가로 55℃까지 역전사 반응이 가능하여 productivity를 증가시켰습니다.

Controllable Reaction Time:
반응시간은 증폭하려는 유전자의 copy 수와 크기에 따라 달라질 수 있습니다. High copy gene의 경우, 10분의 역전사 반응으로도 충분한 양의 cDNA를 합성할 수 있습니다.

Ease-of-Use & Speed: 
순환적 역전사 반응을 수행할 경우 primer와 RNA template의 pre-incubation 과정이 생략되어 실험방법이 간단하고 반응시간이 단축됩니다.

Reproducibility: 
재현성 있는 결과를 위해 바이오니아의 전 제품은 엄격한 ISO 품질 시스템하에서 생산 됩니다.

 

Enzyme properties

5' to 3' exonuclease activity: No
3' to 5' exonuclease activity: No
3' – A overhang: Yes
Strand displacement: Yes
Fragment size: Up to 9 kb


Applications

First-strand synthesis of cDNA from RNA molecules
RT-PCR
Random priming reaction
Library construction
Probe labeling
mRNA 5' end mapping by primer extension analysis


Reagents supplied

5x Reaction Buffer:  Tris-HCl, KCl, MgCl2 (pH8.1)
100 mM DTT
dNTP mix: 2.5 mM of each dNTP


Concentration

10.000 U/50 μl


Storage conditions

50% glycerol containing 20 mM Tris-Cl (pH7.6), 150 mM NaCl, 0.1 mM EDTA, 1 mM DTT, 0.1 % IGEPAL CA-630


Store temperature

-20°C


Unit definition

One unit is defined as the amount of enzyme required to incorporates 1 nmole of dTTP into acid-precipitable material in 10 minutes at 37°C using poly (A) oligo (dT) as template primer.

Experimental Data

figure1

Figure 1. Comparison of transferrin receptor gene amplification with different reverse transcriptases. 700 ng of total RNA was used for reverse transcription and the same amount of amplified products were used for electrophoresis.

Lane MW; 100 bp Plus DNA Ladder (D-1035)
Lane 1 - 4; TFR (Transferrin receptor gene) amplified with MMLV
Lane 5 - 8; TFR amplified with CycleScript
Lane 9 - 12; TFR amplified with CycleScript
Lane 13 - 16; TFR amplified with MMLV from company IA
Lane 17 - 20; TFR amplified with S-script from company S
Lane 21 - 24; TFR amplified with S-script ll from company I
Lane 25 - 28; TFR amplified with S-script lll from company I
Lane 29 - 32; TFR amplified with O-script from company Q


figure2

Figure 2. Comparison of β-actin gene amplification with different reverse transcriptases. Each 10 ng, 1 ng, 100 pg, and 10 pg of total RNA was used for reverse transcription and the same amount of amplified products were used for electrophoresis.

Lane 1 - 4; β-actin amplified with CycleScript
Lane 5 - 8; β-actin amplified with CycleScript
Lane 9 - 12; β-actin amplified with CycleScript
Lane 13 - 16; β-actin amplified with MMLV from company I
Lane 17 - 20; β-actin amplified with S-script from company S
Lane 21 - 24; β-actin amplified with S-script from company S
Lane 25 - 28; β-actin amplified with S-script from company I
Lane 29 - 32; β-actin amplified with S-script from company I


figure3

Figure 3. Comparison of GAPDH gene amplification with different reverse transcriptases. Each 10 ng, 1 ng, 100 pg, and 10 pg of total RNA was used for reverse transcription and the same amount of amplified products were used for electrophoresis.

Lane 1 - 4; GAPDH amplified with CycleScript
Lane 5 - 8; GAPDH amplified with CycleScript
Lane 9 - 12; GAPDH amplified with CycleScript
Lane 13 - 16; GAPDH amplified with MMLV from company I
Lane 17 - 20; GAPDH amplified with S-script from company S
Lane 21 - 24; GAPDH amplified with S-script from company S
Lane 25 - 28; GAPDH amplified with S-script from company I
Lane 29 - 32; GAPDH amplified with S-script from company I


figure4

Figure 4. Working temperature comparison of different reverse transcriptases. Each 10 ng, 1 ng, 100 pg, and 10 pg of total RNA was used for reverse transcription and the same amount of amplified products were used for electrophoresis.

Lane 1 - 4; GAPDH amplified with CycleScript
Lane 5 - 8; GAPDH amplified with CycleScript
Lane 9 - 12; GAPDH amplified with CycleScript
Lane 13 - 16; GAPDH amplified with S-script from company S
Lane 17 - 20; GAPDH amplified with S-script from company S
Lane 21 - 24; GAPDH amplified with S-script from company S


Cat. No. Product Description Price
E-3131 CycleScript Reverse Transcriptase (10,000 U, 50 rxn) ₩84,000
E-3132 CycleScript Reverse Transcriptase (50,000 U, 250 rxn) ₩294,000
EB-1003 5X reaction buffer, 1 ml ₩25,000


















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