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EZ™ T7 High Yield In Vitro Transcription Kit new

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1 EZ™ T7 High Yield In Vitro Transcription Kit EZ027S - 75 reaction ₩ 350,000 추가
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Product description

T7 phage RNA polymerases are widely used for the in vitro synthesis of RNA transcripts from DNA templates.

 Enzynomics EZ™ T7 High Yield In Vitro Transcription Kit is designed for high yield in vitro transcription from DNA templates containing T7 RNA Polymerase promoter. The phage RNA polymerases have a high specificity for their respective promoters. The complete kit includes the RNA polymerases, all of the required reagents for performing transcription reactions in vitro (excluding radioisotope).

 The kit contains sufficient reagents for 75 reactions of 20 μl each. Each standard reaction yields up to 180 μg of RNA from 1 μg control template in 1 hour. The template must have a double-stranded 19–23 base promoter upstream of the sequence to be transcribed. The template is then mixed with the corresponding RNA polymerase, rNTPs, and transcription buffer, and the reaction mixture is incubated for 1 hr at 37°C. RNA polymerase first binds to its double-stranded DNA promoter, then it separates the two DNA strands, and uses the 3' to 5' strand as a template to synthesize a complementary 5' to 3' at the end of the DNA template.

 The kit can be successfully used to produce both long and short RNA transcripts. The RNA synthesized is suitable for variety of applications that require large amounts of RNA, such as in vitro translation, antisense RNA and RNAi studies, RNase protection assays, studies of RNA splicing, isolation of RNA binding proteins. Non-radioactively labeled RNA can be used as probes in microarrays, blots or in situ hybridization.

 

 

Figure 1. The principle of In vitro RNA transcription system.

 

Kits components

- 5X Transcription Buffer

- 10X MgCl2

- rATP (100 mM)

- rGTP (100 mM)

- rCTP (100 mM)

- rUTP (100 mM)

- Control Template DNA (1 μg/μl)

- T7 RNA Polymerase (200 U/μl)

- DTT (100 mM)

20X Enhancer Solution

- Sterile water (RNase free)

 

Storage conditions

All component: -20

 

Additional materials required

- DNase I

- RNA Ladder Marker

- RNA Loading dye

- RNA Loading buffer 

- RNA Sample buffer

 

Note

- The mixture should be kept at room temperature while each successive component is added, since DNA 

  can precipitate in the presence of spermidine if kept at 4℃.

- Avoiding RNase Contamination

- RNA frozen in Transcription buffer, as it will precipitate at low temperatures in the presence of spermidine.

  RNA stored in this way will not run to its true size upon electrophoresis. 

- If the DNA template is not removed, a high-molecular-weight band (~4kb for the control template DNA) 

  will be visible.

- Enhancer Solution is consist of RNase Inhibitor and enzyme cocktail for the stimulation of RNA synthesis

 


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