SuperScript™ III 역전사 효소
SuperScript™ III 역전사 효소
Invitrogen™

SuperScript™ III 역전사 효소

Invitrogen SuperScript III Reverse Transcriptase는 RNase H 활성 감소, 반감기 증가 및 열 안정성 향상을 위해 여러 돌연변이를 도입하여 유전적으로자세히 알아보기
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카탈로그 번호반응 수
1808009310 Reactions
1808004450 Reactions
18080085200 Reactions
카탈로그 번호 18080093
제품 가격(KRW)
150,000
Online offer
Ends: 30-Sep-2025
176,000
할인액 26,000 (15%)
Each
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반응 수:
10 Reactions
대량 주문 또는 맞춤형 요청
제품 가격(KRW)
150,000
Online offer
Ends: 30-Sep-2025
176,000
할인액 26,000 (15%)
Each
카트에 추가하기
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Invitrogen SuperScript III Reverse Transcriptase는 RNase H 활성 감소, 반감기 증가 및 열 안정성 향상을 위해 여러 돌연변이를 도입하여 유전적으로 조작된 MMLV 역전사 효소(RT)입니다. SuperScript III RT는 야생형 MMLV 및 MMLV RNase H-minus 효소보다 더 높은 cDNA 수율, cDNA 길이 개선, GC-rich 표적 RNA에 대한 개선된 효율성 및 전반적으로 더 나은 성능을 제공합니다.

SuperScript RT는 현재까지 50,000 건이 넘는 인용, 리뷰 및 논문과 함께 가장 신뢰성이 높고 널리 사용되는 RT입니다.

참고: SuperScript RT 제품군의 최신 제품인 SuperScript IV Reverse Transcriptase는 최적의 순도 또는 무결성을 제공하며 RNA 샘플에 사용 시 향상된 열안정성, processivity, 수율 및 성능을 제공합니다.

연구용으로만 사용하십시오. 진단용으로는 사용할 수 없습니다.
사양
농도200 U/μL
최종 제품 유형First-Strand cDNA
형식독립형 효소
반응 수10 Reactions
최적 반응 온도50° C
수량2000 유닛
반응 형식Separate components
시약 유형Reverse Transcription
역전사 효소SuperScript™ III
리보뉴클레아제 H 활성감소
배송 조건드라이아이스
크기(최종 제품)Up to 12.3 kb
시작 물질RNA
기술Reverse Transcription
GC-Rich PCR Performance높음
반응 속도표준
Unit SizeEach
구성 및 보관
SuperScript™ III 역전사 효소(총 2,000 유닛, 200U/µL)에는 5X 제1가닥 버퍼[250mM Tris-HCl(pH 8.3), 375mM KCl, 15mM MgCl2] 바이알(1mL) 및 100mM DTT 바이알(500µL)이 제공됩니다. -20°C에서 보관하십시오.
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자주 묻는 질문(FAQ)

Which components of the SuperScript III First Strand Synthesis System for RT-PCR are available for purchase separately?

The following components are available as stand-alone items:

- Superscript III Reverse Transcriptase (Cat. Nos. 18080093, 18080044, 18080085)
- Oligo (dT)20 Primer (Cat. No. 18418020)
- Random hexamers (Cat. No. 48190011)
- 10 mM dNTP Mix (Cat. Nos. 18427013, 18427088)
- RNAseOUT Recombinant Ribonuclease Inhibitor (Cat. No. 10777019)
- E. coli RNAse H (Cat. Nos. 18021014, 18021071)

I am interested in generating cDNA from total RNA. What is the difference between SuperScript III Reverse Transcriptase and SuperScript III First Strand Synthesis System for RT-PCR?

SuperScript III Reverse Transcriptase (Cat. Nos. 18080093, 18080044, 18080085) contains the stand-alone enzyme and a vial each of 5X first-strand buffer and 100 mM DTT.

SuperScript III First Strand Synthesis System for RT-PCR is a complete kit that provides the SuperScript III Reverse Transcriptase and all the other components required for synthesis of first-strand cDNA from total or poly(A)- RNA. It includes:
- Superscript III Reverse Transcriptase
- Oligo (dT)20 Primer
- Random hexamers
- 10X RT buffer
- 25 mM MgCl2
- 0.1 M DTT
- 10 mM dNTP Mix
- RNAseOUT Recombinant Ribonuclease Inhibitor
- E. coli RNAse H
- DEPC-treated water
- Total HeLa RNA control
- Sense control primer
- Anti-sense control primer
Note: The kit does not include the PCR amplification enzyme.

How can I remove genomic DNA contamination from my sample prior to performing RT-PCR?

If amplification products are generated in the control tube/well that contains no reverse transcriptase (i.e., the no-RT control), it may be necessary to eliminate residual genomic DNA from the RNA sample. Use the following protocol to remove genomic DNA from the total RNA preparation.Random primers are the best choice for degraded RNA, RNA with heavy secondary structure, non-polyadenylated RNA, or prokaryotic RNA. It is recommended only for two-step RT-PCR, and typically gives the highest yields, although the cDNA may not necessarily be full length. Oligo(dT) primers are good to use when trying to recover full-length cDNA from 2-step RT-PCR. The reaction is influenced by secondary structure and RNA quality. Gene specific primers should be used for very specific, mainly one-step RT-PCR reactions. Random primers are the best choice for degraded RNA, RNA with heavy secondary structure, non-polyadenylated RNA, or prokaryotic RNA. It is recommended only for two-step RT-PCR, and typically gives the highest yields, although the cDNA may not necessarily be full length. Oligo(dT) primers are good to use when trying to recover full-length cDNA from 2-step RT-PCR. The reaction is influenced by secondary structure and RNA quality. Gene specific primers should be used for very specific, mainly one-step RT-PCR reactions.

Add the following to an autoclaved 0.5 mL microcentrifuge tube on ice:
1.Total RNA, ideally, less than or equal to 1 µg. (See Note 1 below.)
2.1.0 µL of 10X DNase buffer (200 mM Tris, pH 8.3, 500 mM KCl, 20 mM MgCl2).
3.0.1 U-3.0 U of DNase I (RNase-free, Cat. No. 18047019) or 1.0 U Dnase I, Amplification Grade (Cat. No. 18068015. (See Note 2 below.)
4.Bring volume up to 10 µL with DEPC-treated water.
5.Incubate at room temperature for 15 min. (See Note 3 below.)
6.Terminate the reaction by adding 1 µL 25 mM EDTA and heat 10 min at 65 degrees C. (See Note 4 below.)
7.Place on ice for 1 minute.
8.Collect by brief centrifugation. This mixture can be used directly for reverse transcription.

Please note the following:
1.To work with higher quantities of RNA, scale up the entire reaction linearly. Do not exceed 2 µg RNA in the 10 µL reaction. More RNA will increase the viscosity of the solution and prevent the DNAse I from diffusing and finding the DNA.
2.DNAse I, Amplification Grade has been extensively purified to remove trace ribonuclease activities commonly associated with other "RNAse-free" enzyme preparations and does not require the addition of placental RNAse inhibitor.
3.It is important not to exceed the 15 minute incubation time or the room temperature incubation. Higher temperatures and longer times could lead to Mg2+-dependent hydrolysis of the RNA.
4.This procedure requires careful pipetting of all solutions so that the concentration of divalent metal cation (Mg2+) is controlled.
5.Because the DNAse I must be heated to 65 degrees C to inactivate the enzyme, the concentration of free divalent metal ions must be low enough (less than 1 mM) after addition of the EDTA to prevent chemical hydrolysis of the RNA. See references below.
After the addition of EDTA, there is an approximately 1:1 molar ratio of Mg2+ :EDTA. EDTA chelates Mg2+ molecules on a 1:1 molar basis. Therefore, this RNA can be directly used in a reverse transcription reaction. First-strand reverse transcription buffers typically result in a final concentration of 2.5 mM Mg2+. If the reverse transcription buffer does not contain MgCl2, add it to the reaction at a final concentration of 2.5 mM. This results in a net final concentration of approximately 2.25 to 2.5 mM MgCl2.

References on RNA hydrolysis:
Molekulyarnaya Biologiya (1987) 21:1235-1241.
References on the mechanism of hydrolysis by other cations:
Eichorn GL and Butzov JY (1965) Biopolymers 3:79.
Butzov JY and Eichorn GL (1965) Biopolymers 3:95.
Farkas WR (1968) Biochim Biophys Acta 155:401.
The authors of the first paper express the opinion that the mechanism of the nonspecific hydrolysis by cations which proceeds through 2',3' cyclic phosphate formation is similar to that of specific hydrolysis such as RNA splicing.

How much RNA should be employed for first-strand cDNA synthesis?

The amount of RNA template for a cDNA synthesis is highly flexible and depends upon the amount of sample available and an individual's need. In general, 1 µg total RNA is used in a typical 20-µL RT reaction.

Find additional tips, troubleshooting help, and resources within ourReverse Transcription and RACE Support Center.

Should I treat the cDNA with RNase H prior to downstream processing?

Some feel that the RNA in the RNA:DNA duplex after reverse transcription will inhibit PCR primers from annealing and amplifying the cDNA. The RNA is still present when using RNase H-mutant RTs. RNase H frees the cDNA from the RNA. On the other hand, some feel that the 95 degrees C denaturing step will cause the RNA primers to fall off the DNA and therefore RNase H treatment is not necessary. Therefore, this step is optional. For cloning of larger fragments, RNase H treatment can be beneficial.

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