2×HiF Taq faaopoopo Master Mix
Numera pusi: HCR2014B
HIF Taq plus Master Mix (With Dye) o se vaifofo 2×premixed saunia e fa'aoga o lo'o iai le Plus HIF DNA Polymerase, dNTPs, ma fa'apolopolo lelei.E lua monoclonal antibodies i le vevela o le potu e taofia ai le gaioiga o le polymerase ma le 3′→5′exonuclease gaioiga e faʻaopoopo i le faʻafefiloi matai mo le faigofie ma sili ona patino Hot Start PCR.O le faʻaopoopoga faʻaopoopo e faʻaopoopoina i le faʻafefiloi matai e tuʻuina atu ai i le enzyme se malosi faʻalautele vaega umi, o le umi o le faʻaopoopoga e mafai ona oʻo atu i le 13 kb, o le enzyme o loʻo i ai le 5′→3′ DNA polymerase activity ma le 3′→5′. gaoioiga exonuclease, o lona faamaoni e 83 taimi o le Taq DNA polymerase, lea e 9 taimi o le DNA polymerase masani.E fetaui lelei mo le faʻalauteleina o faʻataʻitaʻiga lavelave, o le oloa faʻateleina o se pito faʻamaʻi.
2×HIF Taq plus Master Mix(With Dye) o loʻo i ai le lelei o le vave ma le faigofie, maualuga le lagona, malosi faʻapitoa, lelei le mautu, ma isi mea, o le faiga faʻaalia e manaʻomia naʻo le faʻaopoopoina o primers ma templates, ma e mafai ona faʻalauteleina e se lua- fa'asologa o laasaga, fa'afaigofie laasaga fa'ata'ita'i ma fa'asaoina le taimi.O lenei oloa o lo'o i ai ni vaila'au fa'ailoga electrophoresis, ma o oloa PCR e mafai ona fa'aoga sa'o mo electrophoresis.E le gata i lea, o le oloa foi o loʻo i ai se sui puipuia faʻapitoa, ina ia mafai e le faʻafefiloi matai ona faatumauina le gaioiga mautu pe a uma ona toe faʻafefete.
Tulaga Teuteu
O oloa e tatau ona teuina ile -25~-15℃ mo le 1 tausaga.
Fa'amatalaga
Fa'amatalaga o oloa | Master Mix |
Fa'atotonuga | 2× |
Amata vevela | Amata vevela ua fausia |
Fa'atautau | Tu'u |
Saosaoa tali | Vave |
Tele (Oloa Mulimuli) | E oʻo atu i le 13kb |
Tulaga mo felauaiga | Aisa mago |
Ituaiga oloa | Premixes PCR maualuga fa'amaoni |
Faatonuga
1.PCR Reaction System
Vaega | Volume (μL) |
DNA mamanu | Talafeagai |
Fa'ailoga muamua (10 μmol/L) | 2.5 |
Fa'ata'ita'i (10 μmol/L) | 2.5 |
2×HIF Taq faaopoopo Master Mix | 25 |
ddH2O | i le 50 |
2.Fautuaina le fa'aogaina o mamanu eseese
Ituaiga mamanu | Fa'ateleina vaega mai le 1kb i le 10 kb |
Genomic DNA | 50ng-200 ng |
Plasmid poʻo Viral DNA | 10pg-20ng |
cDNA | 1-2.5 µL (Aua le sili atu i le 10% o le tele o tali PCR mulimuli) |
3.Polokalama Fa'aliga
1) Polokalama Laasaga Lua (fa'ata'ita'iga lavelave)
Laasaga taamilo | Temp. | Taimi | Ta'amilosaga |
Uluai denaturation | 98℃ | 3 minu | 1 |
Denaturation | 98℃ | 10sec | 30-35 |
Fa'aopoopoga | 68℃ | 30 sec/kb | |
Fa'aopoopoga mulimuli | 72 ℃ | 5 minu | 1 |
2) Tolu-Step Protocol (faiga masani)
Laasaga taamilo | Temp. | Taimi | Ta'amilosaga |
Uluai denaturation | 98℃ | 3 minu | 1 |
Denaturation | 98℃ | 10sec | 30-35 |
Fa'asusu | 60℃ | 20 sec | |
Fa'aopoopoga | 72 ℃ | 30 sec/kb | |
Fa'aopoopoga mulimuli | 72 ℃ | 5 minu | 1 |
3) Fa'ata'ita'i Fa'ata'ita'i (Fa'ata'ita'iga lavelave)
Laasaga taamilo | vevela | Taimi | Ta'amilosaga |
Uluai denaturation | 98℃ | 3 minu | 1 |
Denaturation | 98℃ | 10 sec | 15 (1℃ fa'aitiitiga ile ta'amilosaga) |
Fa'alili fa'alelei | 70-55 ℃ | 20 sec | |
Fa'aopoopoga | 72 ℃ | 30 sec/kb | |
Denaturation | 98℃ | 10 sec |
20 |
Fa'asusu | 55℃ | 20 sec | |
Fa'aopoopoga | 72 ℃ | 30 sec/kb | |
Fa'aopoopoga mulimuli | 72 ℃ | 5 minu | 1 |
Fa'ailoga i lalo o fa'asalalauga fa'aopoopo eseese
Protocol | Laasaga Lua | Laasaga tolu | Fa'alili fa'alelei |
Fa'amatalaga. | vave | feololo | telegese |
Fa'apitoa | maualuga | feololo | maualuga |
PCR fua | feololo | maualuga | feololo |
Su'e fua | maualuga | feololo | maualuga |
Fa'amatalaga
Fa'amolemole fai le PPE e mana'omia, o se ofu su'esu'e ma totigilima, ia mautinoa lou soifua maloloina ma le saogalemu!