English name: TDT; Terminal Transferase; Terminal Deoxynucleotidyl Transferase
Other names: terminal deoxynucleotidyl transferase; Thymopentin
Source: E. coli cells containing the coding calf thymus terminal deoxynucleotidyl transferase gene
Concentration: 20u / ul
Activity Definition: refers to the 37 ℃, 60 minutes 1nmol DNA incorporated into a polynucleotide fraction (adsorbed on DE-81) the amount of enzyme required
Activity assay mixture: 66mM potassium cacodylate (PH7.2), 1mM CoCL2, 0.01% (v / v) Triton X-100, 10uM oligo (dT) 10, 1mM dTTP and 0.4MBq / ml [3h] - dTTP
Preservation solution components: 100mM potassium acetate (PH6.8), 2mM β- mercaptoethanol, 0.01% (v / v) Triton X-100 and 50% (v / v) glycerol
5 * reaction buffer: 1M potassium cacodylate, 125mM Tris, 0.05% (v / v) Triton X-1005mM CoCL2 (PH7.225 ℃)
Inhibitors: metal chelators, ammonium, chloride, iodide and phosphate ions
Inactivation: Add EDTA, heated 70 ℃ 10 分钟
Quality assurance: test showed no endo- and exo-DNase, RNase, phosphatase pollution
Note: Due to contain CoCL2, TdT reaction buffer is not compatible with downstream applications, it requires the use of column centrifugation or phenol / chloroform extraction and ethanol precipitation reaction mixture was purified to remove CoCL2
Characters: The suspension recombinase terminal transferase (Terminal transferase, TdT) is a DNA polymerase without template, deoxynucleotide catalytic binding to 3 'hydroxyl end of the DNA molecule with projecting, recessed or blunt ends. . The single-stranded DNA molecule can be used as a substrate for TdT general operation are: first open a single site on the support it with terminal transferase enzyme and a dNTP (eg dATP) incubated together to make Tim tail, and the other one to be inserted exogenous DNA fragment is complementary with the same method used to add dNTP (dTTP) using the same method to add tail. Then make the two are connected to the end of the complementary single-stranded DNA fragments anneal to each other, forming a hybrid vector to transform recipient strain. heterozygous carrier cracks or cuts may be subject to a full restoration by bacteria.
Use: Biochemical studies 3 'end add homopolymers; 3 DNA of the' end of the catalytic DNA markers.
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