BspQI
BspQI le ṣe afihan ni atunbere ni E. coli ti o le ṣe idanimọ awọn aaye kan pato ati ti iṣelọpọ labẹ
BspQI, idinamọ IIs endonuclease ihamọ endonuclease, ti wa lati inu igara E. coli ti o tun ṣe ti o gbe cloned ati iyipada BspQI gen lati Bacillus sphaericus.O le ṣe idanimọ awọn aaye kan pato, ati ilana idanimọ ati awọn aaye fifọ jẹ bi atẹle:
5' · · · · GCTCTTC(N) · · · · · · · · · · · · 3’
3' · · · · CGAGAAG(NNNN) · · · · 5'
Ọja Awọn ẹya ara ẹrọ
1. Iṣẹ-ṣiṣe ti o ga julọ, Tito nkan lẹsẹsẹ;
2. Low star aṣayan iṣẹ-ṣiṣe, aridaju deede gige bi "scalpel";
3. Laisi BSA ati eranko-orisun free;
Ifamọ Methylation
DEmi methylation:Ko Kokoro;
Dcm methylation:Ko Kokoro;
CpG Methylation:Ko Kokoro;
Awọn ipo ipamọ
Ọja naa yẹ ki o firanṣẹ ≤ 0 ℃;Fipamọ ni ipo -25~-15℃.
Ifipamọ ipamọ
20mM Tris-HCl, 0.1mM EDTA, 500 mM KCl, 1.0 mM dithiothreitol, 500 µg/ml Recombinant Albumin, 0. 1% Trition X- 100 ati 50% glycerol (pH 7.0 @ 25°C).
Unit Definition
Ẹyọ kan jẹ asọye bi iye henensiamu ti o nilo lati da 1µg ti DNA iṣakoso inu ni wakati 1 ni 50°C ni iwọn ifasẹyin lapapọ ti 50 µL.
Iṣakoso didara
Ayẹwo Iwa Amuaradagba (SDS-PAGE):Iwa-mimọ ti BspQI jẹ ≥95% ti pinnu nipasẹ itupalẹ SDS-PAGE.
RNase:10U ti BspQI pẹlu 1.6μg MS2 RNA fun awọn wakati 4 ni 50℃ ko ni ibajẹ bi a ti pinnu nipasẹ agarose gel electrophoresis.
Iṣẹ-ṣiṣe DNA ti kii ṣe pato:10U ti BspQI pẹlu 1μg λ DNA ni 50℃ fun wakati 16, ni akawe pẹlu 50℃ fun wakati 1, ko fa DNA ti o pọ ju bi a ti pinnu nipasẹ agarose gel electrophoresis.
Ligation ati didasilẹ:Lẹhin tito nkan lẹsẹsẹ ti 1 μg λDNA pẹlu 10U BspQI, awọn ajẹkù DNA le ni asopọ pẹlu T4 DNA ligase ni 16ºC.Ati pe awọn ajẹkù ligated wọnyi le tun ge pẹlu BspQI.
E. koli DNA: E. coli 16s rDNA-pato TaqMan qPCR wiwa fihan pe E.coli genome iyokù ≤ 0.1pg/ul.
Iyoku amuaradagba:≤50 ppm
Kokoro Endotoxin: LAL-igbeyewo, ni ibamu si Chinese Pharmacopoeia IV 2020 àtúnse, jeli iye igbeyewo ọna, gbogboogbo ofin (1143).Akoonu endotoxin kokoro yẹ ki o jẹ ≤10 EU/mg.
Ifesi eto ati ipo
| Ẹya ara ẹrọ | Iwọn didun |
| BspQ I (10 U/μL) | 1 μL |
| DNA | 1 μg |
| 10 x BspQ Mo ifipamọ | 5 μL |
| dd H2O | Titi di 50 μl |
Awọn ipo idahun: 50 ℃, 1 ~ 16 wakati.
Aiṣiṣẹ igbona:80°C fun iṣẹju 20.
Eto ifaseyin ti a ṣeduro ati awọn ipo le pese ipa tito nkan lẹsẹsẹ henensiamu to dara, eyiti o jẹ fun itọkasi nikan, jọwọ tọka si awọn abajade esiperimenta fun awọn alaye.
Ohun elo ọja
Ihamọ tito nkan lẹsẹsẹ endonuclease, cloning iyara.
Awọn akọsilẹ
1. Awọn iwọn didun ti enzymu ≤ 1/10 ti iwọn didun lenu.
2. Iṣẹ-ṣiṣe irawọ le waye nigbati ifọkansi glycerol jẹ diẹ sii ju 5%.
3. Cleavage aṣayan iṣẹ-ṣiṣe le waye nigbati Sobusitireti ni isalẹ awọn niyanju ratio.
