Hotstart Taq DNA Polymerase
Hot Start Taq DNA Polymerase (Antibody modification) o se DNA polymerase e mafai ona amata vevela mai le Thermus aquaticus YT-1, o loʻo iai se gaioiga 5′→3′ polymerase ma le 5' flap endonuclease.O le Taq DNA polymerase vevela-amata o le Taq DNA polymerase lea na suia e le thermolabile Taq antibodies.O le suiga o le antibody na fa'atuputeleina ai le fa'apitoa, lagona ma'ale'ale, ma le maua mai o le PCR.
Vaega
| Vaega | HC1012A-01 | HC1012A-02 | HC1012A-03 | HC1012A-04 |
| 5 × HC Taq Faʻamau | 4×1 mL | 4×10 mL | 4×50 mL | 5×400 mL |
| Hot Start Taq DNA Polymerase (Suia le Antibody) (5 U/μL) | 0.1 mL | 1 mL | 5 ml | 10×5 mL |
Talosaga
10 mM Tris-HCl (pH 7.4 i le 25 ℃), 100 mM KCl, 0.1 mM EDTA, 1 mM dithiothreitol, 0.5% Tween20, 0.5% IGEPALCA-630 ma 50% Glycerol.
Tulaga Teuteu
Felauaiga i lalo ole 0°C ma ia teuina ile -25°C~-15°C.
Fa'amatalaga Vaega
O le tasi iunite o lo'o fa'amatalaina o le aofa'i o le enzyme e fa'aofia ai le 15 nmol o le dNTP i mea e le mafai ona fa'asalaina i totonu ole 30 minute ile 75°C.
Puleaina lelei
1.EndGaoioiga onuclease:O le incubation o le 20 U o le enzyme ma le 4 μg pUC19 DNA mo le 4 itula i le 37 ℃ na iʻu ai i le leai o se faʻaleagaina o le DNA e pei ona fuafuaina e gel electrophoresis.
2.5 kb Lambda PCR:25 Ta'amilosaga o le PCR fa'ateleina o le 5 ng Lambda DNA ma 1.25 iunite o le Taq DNA Polymerase i le i ai o 200 µM dNTPs ma 0.2 µM primers e maua ai le 5 kb oloa fa'amoemoeina.
3.Gaoioiga Exonuclease:O le incubation o le 50 µl tali o lo'o i ai le itiiti ifo o le 12.5 U o le Taq DNA Polymerase fa'atasi ai ma le 10 nmol 5'-FAM oligonucleotide mo le 30 minute i le 37 ℃ e le maua ai se fa'aleagaina.
4.RNase Gaoioiga:O le incubation o le 10 μL faʻaalia o loʻo i ai le 20 U o le enzyme ma le 1μg o RNA transcripts mo le 2 itula i le 37 ° C na mafua ai le leai o se faʻaleagaina o le RNA e pei ona fuafuaina e gel electrophoresis.
5.Fa'aleaogaina o le vevela:Leai.
Reaction System
| Vaega | Volume |
| DNA mamanua | faitalia |
| 10 μM Ulua'i Muamua | 0.5 μL |
| 10 μM Fa'aliliuga Muamua | 0.5 μL |
| dNTP Mix (10mM ta'itasi) | 0.5 μL |
| 5 × HC Taq Faʻamau | 5 μL |
| Taq DNA Polymerasee(5U/μL) | 0.125 μL |
| Vai e leai ni nuclease | E oʻo atu i le 25 μL |
Fa'amatalaga:
1) a.
| DNA | Aofaiga |
| Genomic | 1 ng-1 μg |
| Plasmid poʻo Viral | 1 itulau-1 ng |
2) e.Ole fa'atonuga sili ona lelei ole Taq DNA Polymerase e mafai ona maua mai i le 5-50 units/mL (0.1-0.5 units/25 μL reaction) i fa'aoga fa'apitoa.
Polokalama uila uila vevela
PCR
| Laa | vevela(°C) | Taimi | Ta'amilosaga |
| Uluai denaturationa | 95 ℃ | 1-3min | - |
| Denaturation | 95 ℃ | 15-30 s | 30-35 Ta'amilosaga |
| Fa'asusue | 45-68 ℃ | 15-60 s | |
| Fa'aopoopoga | 68 ℃ | 1kb/min | |
| Fa'aopoopo Mulimuli | 68 ℃ | 5min | - |
Fa'amatalaga:
1) O le faʻauluina muamua o le 1 min i le 95 ° C e lava mo le tele o faʻaopoopoga.Mo faʻataʻitaʻiga faigata, e fautuaina le umi o le denaturation o le 2-3mins i le 95 ° C.Fa'atasi ai ma colony PCR, e fautuaina le fa'amalo ole 5mins muamua ile 95°C.
2) O le laasaga faʻapipiʻi e masani lava 15-60 s.E fa'atatau le vevela o le su'ega ile Tm ole pa'aga muamua ma e masani lava ole 45-68℃.
