Warm Start Bst 2.0 DNA Polymerase (Glycerol yaulere)
Bst DNA polymerase V2 imachokera ku Bacillus stearothermophilus DNA Polymerase I, yomwe ili ndi 5'→ 3' DNA polymerase ntchito ndi ntchito yamphamvu yowonjezera unyolo, koma palibe 5′→3′ exonuclease ntchito.Bst DNA Polymerase V2 ndiyoyenera kusamuka kwa strand, isothermal amplification LAMP (Loop mediated isothermal amplification) ndikutsatizana mwachangu.Bst DNA polymerase V2 ndi mtundu woyambira wotentha wotengera Bst DNA polymerase V2 (HC5005A) wopezedwa ndiukadaulo wosinthika wosinthika, womwe ungalepheretse ntchito ya DNA polymerase kutentha kwa firiji, kotero kuti makina amachitidwe amatha kuyendetsedwa ndikupangidwira kutentha kwachipinda kuti apewe -kukulitsa kwapadera ndikuwongolera magwiridwe antchito, ndipo bukuli limatha kukhala lyophilized.Kuonjezera apo, ntchito yake imatulutsidwa pa kutentha kwakukulu, kotero palibe chifukwa cha sitepe yotsegulira.
Zigawo
| Chigawo | Mtengo wa HC5006A-01 | Mtengo wa HC5006A-02 | Mtengo wa HC5006A-03 |
Bst DNA polymerase V2 (Glycerol-free) (8U/μL) | 0.2 ml pa | 1 ml | 10 ml pa |
| 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 |
Mapulogalamu
1.LAMP isothermal amplification
2.DNA strand single displacement reaction
3.High GC gene kutsatizana
4.Kutsata kwa DNA kwa nanogram level.
Mkhalidwe Wosungira
Kuyenda pansi pa 0 ° C ndikusungidwa pa -25 ° C ~ -15 ° C.
Tanthauzo la Chigawo
Chigawo chimodzi chimatanthauzidwa ngati kuchuluka kwa enzyme yomwe imaphatikiza 25 nmol ya dNTP muzinthu zosasungunuka za asidi mu mphindi 30 pa 65 ° C.
Kuwongolera Kwabwino
1.Protein Purity Assay (SDS-PAGE):Kuyera kwa Bst DNA polymerase V2 ndi ≥99% yotsimikiziridwa ndi kusanthula kwa SDS-PAGE pogwiritsa ntchito kuzindikira kwa Coomassie Blue.
2.EndonucleaseZochita:Kuyika kwa 50 μL reaction yokhala ndi osachepera 8 U ya Bst DNA polymerase V2 yokhala ndi 1 μg λDNA kwa maola 16 pa 37 ℃ kumapangitsa kuti pasakhale kuwonongeka komwe kumadziwika.
3.Ntchito ya Exonuclease:Kuyika kwa 50 μL reaction yokhala ndi osachepera 8 U ya Bst DNA polymerase V2 yokhala ndi 1 μg λ -Hind Ⅲ digest DNA kwa maola 16 pa 37 ℃ kumapangitsa kuti pasakhale kuwonongeka komwe kwatsimikiziridwa.
4.Ntchito ya Nickase:Kuyika kwa 50 μL reaction yokhala ndi osachepera 8 U ya Bst DNA polymerase V2 yokhala ndi 1 μg pBR322 DNA kwa maola 16 pa 37°C kumapangitsa kuti pasakhale kuwonongeka kodziwika monga momwe zatsimikizidwira.
5.Ntchito ya RNase:Kuyika kwa 50 μL reaction yokhala ndi osachepera 8 U ya Bst DNA polymerase V2 yokhala ndi 1.6 μg MS2 RNA kwa maola 16 pa 37°C kumapangitsa kuti pasakhale kuwonongeka kodziwika monga momwe zatsimikizidwira.
6.E. koliDNA:120 U of Bst DNA polymerase V2 imawunikiridwa ngati pali E. coli genomic DNA pogwiritsa ntchito TaqMan qPCR yokhala ndi zoyambira za E. coli 16S rRNA locus.Kuwonongeka kwa E. coli genomic DNA ndi ≤1 Copy.
Kusintha kwa LAMP
| Zigawo | 25μL |
| 10 × HC Bst V2 Buffer | 2.5 μL |
| MgSO4 (100mM) | 1.5 μL |
| dNTPs (10mM iliyonse) | 3.5 μl |
| SYTO™ 16 Green (25×)a | 1.0 μL |
| Kusakaniza koyambirirab | 6 ml |
| Bst DNA Polymerase V2 (Glycerol-free) (8 U/uL) | 1 ml |
| Template | × μl ndi |
| ddH₂O | Mpaka 25 μL |
Ndemanga:
1) a.SYTOTM 16 Green (25 ×): Malingana ndi zosowa zoyesera, utoto wina ukhoza kugwiritsidwa ntchito m'malo;
2) b.Kusakaniza koyamba: kupezedwa posakaniza 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 ndi mabuku ena.
Zochita ndi Mkhalidwe
1 × HC Bst V2 Buffer, kutentha kwa ma incubation kuli pakati pa 60°C ndi 65°C.
Kutentha Kutentha
80 ° C, mphindi 20
