T4 RNA Ligase 1,specification,price,image-Bio-Equip in China

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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. [2] Tessier, Daniel C. , R. Brousseau , and T. Vernet . "Ligation of single-stranded oligodeoxyribonucleotides by T4 RNA ligase." Analytical Biochemistry 158.1(1986):171-178.

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

One unit is defined as the amount of enzyme required to convert 1 nanomole of 5´-[32P]rA16 into a phosphatase-resistant form in 30 minutes at 37°C.