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Description
T4 RNA Ligase 1Product Specification Synonyms T4 RNA Ligase, T4 Rnl1 Expression System E. coli Molecular Weight 69kDa (Reducing) Purity 95% by SDS PAGE Tag His Tag Storage Buffer 10 mM Tris HCl50 mM KCl1 mM DTT0. 1 mM EDTA50% Glycerol pH 7. 4 @ 25C Stability & Storage Store at 25 ~ 15 for 2 years Reference [1] England, Thomas E , and O. C. Uhlenbeck . "3 [prime] Terminal labelling of RNA with T4 RNA ligase." Nature 275. 5680(1978): 560 1. [2] Tessier, Daniel C. , R.
Product Specification
| Synonyms | T4 RNA Ligase, T4 Rnl1 |
| Expression System | E.coli |
| Molecular Weight | 69kDa (Reducing) |
| Purity | ＞95% by SDS-PAGE |
| Tag | His Tag |
| Storage Buffer | 10 mM Tris-HCl、50 mM KCl、1 mM DTT、0.1 mM EDTA、50% Glycerol pH 7.4 @ 25°C |
| Stability & Storage | Store at -25 ~ -15℃ for 2 years |
| Reference | [1] England, Thomas E , and O. C. Uhlenbeck . "3|[prime]|-Terminal labelling of RNA with T4 RNA ligase." Nature 275.5680(1978):560-1. |
Background
T4 RNA Ligase 1 is an ATP-dependent enzyme that catalyzes the formation of phosphodiester bonds between the 5'-P and 3'-OH ends of single-stranded RNA, single-stranded DNA, or single nucleotides, either intermolecular or intramolecular. It is suitable for the connection of single-stranded RNA molecules and the 5' end junction constructed by miRNA libraries in NGS. Labeled with RNA 3 'terminal; Cyclization of oligonucleotides; tRNA modification; In 5'-RACE (Rapid Amplification of cDNA Ends) assay, it is used for oligonucleotides to bind to single-stranded cDNA; and the introduction of unnatural amino acids into proteins.
Components
Name |
Storage Temperature |
quantity |
concentration |
T4 RNA Ligase 1 |
-20 ℃ |
100 ul or 500 µl |
10 U/µl |
10*Reaction Buffer |
-20 ℃ |
1 ml |
10 X |
Adenosine 5'-Triphosphate (ATP) |
-20 ℃ |
1 ml |
10 mM |
PEG8000 (RNase free) |
-20 ℃ |
1 ml |
50% |
Protocol
1. Set up a 20 μl reaction as follows:
Reagent |
Volume |
Final Concentration |
10 * Reaction Buffer |
2 ul |
1 X |
Single-stranded RNA with 5´P and 3´OH ends (200 ng-1 µg) |
1 ul |
10-50 ng/ul |
50% PEG8000 |
4 ul |
10% |
1mM ATP |
1 ul |
50 µM |
40 μ/ul Rnase inhibitor |
0.5ul |
1 U/µl |
10 U/µl T4 RNA Ligase 1 |
1ul |
0.5 U/µl |
DEPC-treated Water |
Up to 20ul |
- |
2. Incubate at 37°C for 1-2 hours.
For longer oligos, overnight incubation at 16°C for 16h may improve yield.
3. Incubate at 65°C for 15min to terminate the reaction.
Guidelines
Please avoid repeated freeze-thaw cycles.
Unit Definition
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