Wild Taq DNA Polymerase
Taq DNA Polymerase shine DNA polymerase mai zafi daga Thermus aquaticus YT-1, wanda ke da aikin 5′→3′ polymerase da 5′ flap endonuclease act.
Abubuwan da aka gyara
| Bangaren | HC1010A-01 | HC1010A-02 | HC1010A-03 | HC1010A-04 |
| 10× Taq Buffer | 2 × 1 ml | 2 × 10 ml | 2 × 50 ml | 5 × 200 ml |
| Taq DNA Polymerase (5 U/μL) | 0.1 ml | 1 ml | ml 5 | 5 × 10 ml |
Yanayin Ajiya
Jirgin da ke ƙarƙashin 0 ° C kuma a adana shi a -25 ° C ~ -15 ° C.
Ma'anar Naúrar
An ayyana raka'a ɗaya azaman adadin enzyme wanda ya haɗa 15 nmol na dNTP cikin abu maras narkewa a cikin mintuna 30 a 75°C.
Kula da inganci
1.Gwajin Tsaftar Protein (SDS-PAGE):Tsaftar Taq DNA polymerase an ƙaddara ≥95% ta binciken SDS-PAGE.
2.EndAyyukan onuclease:Aƙalla 5 U na Taq DNA polymerase tare da 1 μg λDNA na tsawon awanni 16 a 37 ℃ yana haifar da rashin lalacewa da za a iya ganowa kamar yadda aka ƙaddara.
3.Ayyukan Exonuclease:Aƙalla 5 U na Taq DNA polymerase tare da 1 μg λ -Hind Ⅲ narke DNA na sa'o'i 16 a 37 ℃ yana haifar da rashin lalacewa da za a iya ganowa kamar yadda aka ƙaddara.
4.Ayyukan Nickase:Matsakaicin 5 U na Taq DNA polymerase tare da 1 μg pBR322 DNA na tsawon awanni 16 a 37°C yana haifar da rashin lahani da za a iya ganowa kamar yadda aka ƙaddara.
5.Ayyukan RNase:Matsakaicin 5 U na Taq DNA polymerase tare da 1.6 μg MS2 RNA na awanni 16 a 37°C yana haifar da rashin lahani da za a iya ganowa kamar yadda aka ƙaddara.
6.E. coliDNA:5 U na Taq DNA polymerase an duba shi don kasancewar E. coli genomic DNA ta amfani da TaqMan qPCR tare da abubuwan da aka keɓance don wurin E. coli 16S rRNA.Kwafin halittar E. coli na DNA shine ≤1 Kwafi.
7.PCR Amplification (5.0kb Lambda DNA)- Halin 50 µL mai ɗauke da 5 ng Lambda DNA tare da raka'a 5 na Taq DNA Polymerase don zagayowar 25 na sakamakon haɓaka PCR a cikin samfurin 5.0kb da ake tsammanin.
Saitin Amsa
| Abubuwan da aka gyara | Ƙarar |
| Samfurin DNAa | na zaɓi |
| 10 μM Gabatarwa | 1 μl |
| 10 μM Reverse Primer | 1 μl |
| dNTP Mix (10mM kowane) | 1 μl |
| 10× Taq Buffer | 5 μl |
| Taq DNA Polymeraseb | 0.25 ml |
| Ruwa mara nukiliya | Har zuwa 50 μl |
Bayanan kula:
1) A mafi kyau duka dauki taro na daban-daban shaci ne daban-daban.Tebu mai zuwa yana nuna shawarar samfurin amfani da tsarin amsawa na 50 µL.
| DNA | Adadin |
| Genomic | 1 ng-1 μg |
| Plasmid ko Viral | 1 shafi-1 ng |
2) Madaidaicin taro na Taq DNA Polymerase na iya zuwa daga 0.25 µL ~ 1 µL a cikin aikace-aikace na musamman.
MartaniShirin
| Mataki | Zazzabi(°C) | Lokaci | Zagaye |
| denaturation na farkoa | 95 ℃ | 5 min | - |
| Denaturation | 95 ℃ | 15-30 s | Zagaye 30-35 |
| Annealingb | 60 ℃ | 15 s ku | |
| Tsawaita | 72 ℃ | 1kb/min | |
| Ƙarshe Ƙarshe | 72 ℃ | 5 min | - |
Bayanan kula:
1) Yanayin denaturation na farko ya dace da yawancin halayen haɓakawa kuma za'a iya gyara su bisa ga ƙayyadaddun tsarin samfuri.Idan tsarin samfuri yana da rikitarwa, za a iya ƙara lokacin pre-denaturation zuwa 5 – 10mins don inganta tasirin ƙima na farko.
2) The annealing zafin jiki yana bukatar a gyara bisa ga Tm darajar na firamare, wanda aka kullum saita zuwa 3 ~ 5 ℃ kasa da Tm darajar na farko;Don hadaddun samfura, wajibi ne a daidaita zafin jiki na annealing da tsawaita lokacin haɓaka don cimma ingantaccen haɓakawa.
