Isiqalo esifudumeleyo Bst 2.0 DNA Polymerase (iGlycerol yasimahla)
I-Bst DNA polymerase V2 ivela kwi-Bacillus stearothermophilus DNA Polymerase I, enomsebenzi we-5′→3′ DNA polymerase kunye nomsebenzi oqinileyo wokutshintshwa kwekhonkco, kodwa akukho 5′→3′ umsebenzi we-exonuclease.I-Bst DNA Polymerase V2 ifanelekile ngokufanelekileyo kwi-strand-displacement, isothermal amplification LAMP (i-Loop mediated isothermal amplification) kunye nokulandelelana ngokukhawuleza.I-Bst DNA polymerase V2 luguqulelo olushushu olusekwe kwi-Bst DNA polymerase V2 (HC5005A) efunyenwe ngetekhnoloji yokuguqula ukuguqulwa, enokuthintela umsebenzi we-DNA polymerase kwiqondo lobushushu begumbi, ngoko ke inkqubo yokusabela inokuqhutywa kwaye yenziwe kwiqondo lobushushu begumbi ukuthintela -i-amplification ethile kunye nokuphucula ukusebenza kakuhle, kwaye le nguqulo ingaba lyophilized.Ukongezelela, umsebenzi wayo ukhululwe kumaqondo aphezulu, ngoko akukho mfuneko yesinyathelo sokuvula esahlukileyo.
Amacandelo
| Icandelo | HC5006A-01 | HC5006A-02 | HC5006A-03 |
I-Bst DNA polymerase V2 (iGlycerol-free) (8U/μL) | 0.2 ml | 1 ml | 10 ml |
| I-10 × HC Bst V2 Buffer | 1.5 ml | 2×1.5 mL | 3×10 mL |
| MgSO4(100mM) | 1.5 ml | 2×1.5 mL | 2×10 mL |
Usetyenziso
1.I-LAMP isothermal amplification
2.I-DNA strand single displacement reaction
3.Ulandelelwano lwemfuza yeGC ephezulu
4.Ukulandelelana kwe-DNA kwinqanaba le-nanogram.
Imeko yoGcino
Ukuthutha ngaphantsi kwe-0°C kwaye igcinwe kwi-25°C~-15°C.
Inkcazelo yeyunithi
Iyunithi enye ichazwa njengobungakanani be-enzyme edibanisa i-25 nmol ye-dNTP kwizinto ezingenayo i-asidi kwimizuzu engama-30 kwi-65 ° C.
Ulawulo lwemeko
1.Uvavanyo lweProtein Purity Assay (SDS-PAGE):Ukucoceka kwe-Bst DNA polymerase V2 ≥99% igqitywe luhlalutyo lwe-SDS-PAGE usebenzisa i-Coomassie Blue ubhaqo.
2.EndonyukliyaUmsebenzi:Ukufukanyelwa kwe-50 μL reaction equlathe ubuncinci be-8 U ye-Bst DNA polymerase V2 ene-1 μg λDNA kwiiyure ezili-16 ngama-37 ℃ ibangela ukuba kungabikho kuthotywa okubonakalayo njengoko kumisiwe.
3.Umsebenzi we-Exonuclease:Ukufukanyelwa kwe-50 μL reaction equlathe ubuncinci be-8 U ye-Bst DNA polymerase V2 ene-1 μg λ -Hind Ⅲ yokwetyisa i-DNA iiyure ezili-16 ngama-37 ℃ kubangela ukuba kungabikho kuthotywa kubonwa njengoko kumisiwe.
4.Umsebenzi we-Nickase:Ukufukanyelwa kwe-50 μL reaction equlathe ubuncinci be-8 U ye-Bst DNA polymerase V2 ene-1 μg pBR322 DNA iiyure ezili-16 kuma-37°C ibangela ukuba kungabikho kuthotywa kubonwa njengoko kumisiwe.
5.Umsebenzi we-RNase:Ukufukanyelwa kwe-50 μL reaction equlathe ubuncinci be-8 U ye-Bst DNA polymerase V2 ene-1.6 μg MS2 RNA ngeeyure ezili-16 kuma-37°C ibangela ukuba kungabikho kuthotywa kubonwa njengoko kumisiwe.
6.E. coliI-DNA:I-120 U ye-Bst DNA polymerase V2 ihlolwe ubukho be-E. coli genomic DNA kusetyenziswa i-TaqMan qPCR kunye ne-primers ekhethekileyo ye-E. coli 16S rRNA locus.Ungcoliseko lwe-E. coli genomic DNA yi ≤1 Ikopi.
Isibane Reaction
| Amacandelo | 25μL |
| I-10 × HC Bst V2 Buffer | 2.5 μL |
| MgSO4 (100mM) | 1.5 μL |
| dNTPs (10mM nganye) | 3.5 μL |
| I-SYTO™ 16 eluhlaza (25×)a | 1.0 μL |
| I-Primer mixb | 6 μL |
| I-Bst DNA Polymerase V2 (iGlycerol-free) (8 U/uL) | 1 μL |
| Isifanekiso | × μL |
| ddH₂O | Ukuya kuthi ga kwi-25 μL |
Amanqaku:
1) a.I-SYTO TM 16 eluhlaza (25 ×): Ngokweemfuno zovavanyo, ezinye iidayi zingasetyenziswa njengeendawo;
2) b.I-Primer mix: ifunyenwe ngokuxuba i-20 µ M FIP, 20 µ M BIP, 2.5 µ M F3, 2.5 µ M B3, 5 µ M LF, 5 µ M LB kunye neminye imiqulu.
Ukusabela kunye neMeko
1 × HC Bst V2 Buffer, ubushushu bokufukamela buphakathi kwama-60°C kunye nama-65°C.
Ukushisa Ukungasebenzi
80°C, imizuzu engama-20
