BspQI
BspQI tuaj yeem rov hais dua hauv E. coli uas tuaj yeem paub txog cov chaw tshwj xeeb thiab raug tsim nyob rau hauv
BspQI, ib qho IIs txwv endonuclease txwv endonuclease, yog muab los ntawm ib tug recombinant E. coli hom uas nqa cov cloned thiab hloov BspQI gen los ntawm Bacillus sphaericus.Nws tuaj yeem lees paub qhov chaw tshwj xeeb, thiab kev lees paub ua ntu zus thiab qhov chaw tawg yog raws li hauv qab no:
5' · · · · · · GCTCTTC(N) · · · · · · · · · · · · · · · ·
3' · · · · · CGAGAAG(NNNN) · · · · · 5'
Khoom nta
1. Kev ua haujlwm siab, kev zom zaub mov sai;
2. Tsawg hnub qub kev ua si, xyuas kom raug txiav zoo li "scalpel";
3. Tsis muaj BSA thiab tsiaj keeb kwm dawb;
Methylation Sensitivity
Dkuv methylation:Tsis rhiab heev;
Dcm methylation:Tsis rhiab heev;
CpG Methylation:Tsis rhiab heev;
Cia tej yam kev mob
Cov khoom yuav tsum tau xa ≤ 0 ℃;Khaws ntawm -25 ~ - 15 ℃ mob.
Cia tsis
20mM Tris-HCl, 0.1mM EDTA, 500 mM KCl, 1.0 mM dithiothreitol, 500 µg/ml Recombinant Albumin, 0. 1% Trition X-100 thiab 50% glycerol (pH 7.0 @ 25 ° C).
Unit txhais
Ib chav tsev yog txhais raws li tus nqi ntawm cov enzyme uas yuav tsum tau zom 1µg ntawm Internal tswj DNA hauv 1 teev ntawm 50 ° C hauv tag nrho cov tshuaj tiv thaiv ntim ntawm 50 µL.
Kev Tswj Xyuas Zoo
Protein Purity Assay (SDS-PAGE):Lub purity ntawm BspQI yog ≥95% txiav txim los ntawm SDS-PAGE tsom xam.
RNase:10U ntawm BspQI nrog 1.6μg MS2 RNA rau 4 teev ntawm 50 ℃ yields tsis degradation raws li txiav txim los ntawm agarose gel electrophoresis.
Tsis-Specific DNase Activity:10U ntawm BspQI nrog 1μg λ DNA ntawm 50 ℃ rau 16 teev, piv nrog 50 ℃ rau 1 teev, yields tsis dhau DNA raws li txiav txim los ntawm agarose gel electrophoresis.
Ligation thiab Recutting:Tom qab kev zom ntawm 1 μg λDNA nrog 10U BspQI, DNA fragments tuaj yeem ligated nrog T4 DNA ligase ntawm 16ºC.Thiab cov tawg tawg no tuaj yeem raug txiav nrog BspQI.
E. coli DNA: E. coli 16s rDNA-specific TaqMan qPCR kuaj pom tias E.coli genome residue ≤ 0.1pg/ul.
Tus tswv tsev protein residue:≤ 50 ppm
Kab mob Endotoxin: LAL-test, raws li Suav Pharmacopoeia IV 2020 ib tsab, gel limit test method, general rule (1143).Cov kab mob endotoxin cov ntsiab lus yuav tsum yog ≤10 EU / mg.
Cov tshuaj tiv thaiv kab mob thiab cov xwm txheej
| Cheebtsam | Ntim |
| BspQ I (10 U / μL) | 1 mL ib |
| DNA | 1m g ib |
| 10 x BspQ kuv Buffer | 5l miv |
| dd H2 ua | Txog li 50 μL |
Cov kev mob tshwm sim: 50 ℃, 1 ~ 16 hnub.
Kub inactivation: 80 ° C rau 20 min.
Kev pom zoo cov tshuaj tiv thaiv thiab cov xwm txheej tuaj yeem muab cov txiaj ntsig zoo enzyme digestion, uas yog rau kev siv nkaus xwb, thov xa mus rau qhov kev sim ua kom paub meej.
Daim ntawv thov khoom
Kev txwv endonuclease digestion, ceev cloning.
Sau ntawv
1. Lub ntim ntawm enzyme ≤ 1/10 ntawm cov tshuaj tiv thaiv ntim.
2. Lub hnub qub kev ua si yuav tshwm sim thaum glycerol concentration ntau dua 5%.
3. Cleavage kev ua si yuav tshwm sim thaum Substrate hauv qab qhov piv txwv pom zoo.
