T4 RNA Ligase

English name：T4 RNA Ligase
Molecular weight：43.6kDa，Monomer

Grade：BR
Source: E. coli containing T4 phage clone gene 63

Concentration：10u/ul
Activity definition：It refers to the amount of enzyme required to convert 1nmol 5 '- [32P] - (a) 12-18 to phosphatase resistant form within 30 minutes at 37 ℃.

Enzyme activity analysis mixture：50mM Tris-HCL(PH7.5), 10mM DTT, 10mM MgCL2, 1mM ATP, 10uM 5'-[32P]-(A)12-18(10uM 5'-end）
Preservation solution components：20mM Tris-HCL(PH7.5), 50mM KC1, 1mM DTT，0.1mM EDTA，0.5mM ELUGENT  Denaturant and 50% (v/v) glycerol10×Reaction buffer：500mM HEPES-NaOH(PH8.0，25℃), 100mM MgCL2, 100mM DTT
Inhibitors: Metal chelating agent, SH group modifying reagent

Inactivation: Heat at 70 ℃ for 10 minutes.

Quality control: The test shows that there is no internal and external deoxyribonuclease, ribonuclease and phosphatase pollution.

Note: The concentration of ATP in the reaction system depends on the type of connection reaction, and the recommended concentration of BSA reaction is 0.1mg/ml.

Physicochemical properties: Recombinase catalyzes ATP dependent intramolecular and intermolecular phosphate diester bond connection (5 '- phosphate group and 3' - hydroxyl terminal). The substrate molecules are polynucleotide, oligonucleotide, ssRNA and ssDNA. The smallest connecting substrate between molecules is nucleoside 3 ', 5' - diphosphate, while the smallest substrate connected within molecules is 8-base oligonucleotide.Provide reaction buffer, ATP and BSA solution.It catalyzes the formation of 3 '→ 5' phosphate diester bond and connects the 5 '- phosphate end and 5' - hydroxyl end of nucleotide. The substrates include single stranded RNA, DNA and dinucleotide pyrophosphate.10x reaction buffer was provided with the enzyme.

Purpose: biochemical research. Connecting RNA and RNA; RNA3 '- terminal labeled cytosine 3', 5 '- bis [a-32p] phosphate; Specific modification of tRNA;Cyclization of synthetic oligonucleotides; Oligodeoxyribonucleotides were linked to single stranded cDNA for 5'RACE (rapid amplification of cDNA ends) analysis.