BspQI
Ana iya sake bayyana BspQI a cikin E. coli wanda zai iya gane takamaiman rukunin yanar gizo kuma ana samarwa a ƙarƙashin
BspQI, an IIs ƙuntatawa endonuclease ƙuntatawa endonuclease, an samo shi daga recombinant E. coli iri wanda ke ɗauke da cloned da kuma gyara BspQI gen daga Bacillus sphaericus.Yana iya gane takamaiman rukunin yanar gizo, kuma jerin abubuwan ganowa da wuraren tsagewa sune kamar haka:
5' · · · · GCTCTTC(N) · · · · · · · · · · 3'
3' · · · · CGAGAAG(NNNN) · · · 5'
Siffofin Samfur
1. Babban aiki, saurin narkewa;
2. Low star aiki, tabbatar da daidai yanke kamar "scalpel";
3. Ba tare da BSA da dabba ba kyauta;
Methylation Sensitivity
Dmethylation:Ba Hankali;
Dcm methylation:Ba Hankali;
CpG Methylation:Ba Hankali;
Yanayin ajiya
Ya kamata a aika samfurin ≤ 0 ℃;Adana a yanayin -25 ~ 15 ℃.
Ma'ajiyar ajiya
20mM Tris-HCl, 0.1mM EDTA, 500 mM KCl, 1.0 mM dithiothreitol, 500 μg/ml Recombinant Albumin, 0. 1% Trition X- 100 da 50% glycerol (pH 7.0 @ 25°C).
Ma'anar Naúrar
An bayyana raka'a ɗaya azaman adadin enzyme da ake buƙata don narkar da 1µg na DNA na ciki a cikin awa 1 a 50 ° C a cikin jimlar amsawar 50 µL.
Kula da inganci
Gwajin Tsaftar Protein (SDS-PAGE):Tsaftar BspQI an ƙaddara ≥95% ta binciken SDS-PAGE.
RNase:10U na BspQI tare da 1.6μg MS2 RNA na tsawon awanni 4 a 50℃ ba ya haifar da lalacewa kamar yadda aka ƙaddara ta agarose gel electrophoresis.
Ayyukan DNA marasa Takamaiman:10U na BspQI tare da 1μg λ DNA a 50 ℃ na 16 hours, idan aka kwatanta da 50 ℃ na 1hour, ba ya haifar da wuce haddi DNA kamar yadda aka ƙaddara ta agarose gel electrophoresis.
Ragewa da Gyara:Bayan narkewar 1 μg λDNA tare da 10U BspQI, ana iya haɗa gutsuttsura DNA tare da T4 DNA ligase a 16ºC.Kuma ana iya yanke waɗannan gutsuttsuran ligated tare da BspQI.
E. coli DNA: E.coli 16s rDNA takamaiman TaqMan qPCR ganowa ya nuna cewa ragowar E.coli genome ≤ 0.1pg/ul.
Ragowar furotin mai masauki:≤ 50 ppm
Kwayoyin cuta Endotoxin: LAL-gwajin, bisa ga Pharmacopoeia IV 2020 na Sinanci, hanyar gwajin iyaka ta gel, ƙa'ida ta gaba ɗaya (1143).Ya kamata abun cikin endotoxin na ƙwayoyin cuta ya zama ≤10 EU/mg.
Tsarin amsawa da yanayi
| Bangaren | Ƙarar |
| BspQ I (10 U/μL) | 1 μl |
| DNA | 1 μg |
| 10 x BspQ I Buffer | 5 μl |
| dd H2O | Har zuwa 50 μl |
Yanayin amsawa: 50 ℃, 1 ~ 16 h.
Rashin kunna zafi:80°C na mintuna 20.
Tsarin halayen da aka ba da shawarar da yanayi na iya samar da ingantaccen sakamako mai narkewar enzyme, wanda shine don tunani kawai, da fatan za a koma ga sakamakon gwaji don cikakkun bayanai.
Aikace-aikacen samfur
Ƙuntataccen narkewar endonuclease, cloning mai sauri.
Bayanan kula
1. Girman enzyme ≤ 1/10 na ƙarar amsawa.
2. Ayyukan tauraro na iya faruwa lokacin da ƙwayar glycerol ya fi 5%.
3. Ayyukan ƙwanƙwasa na iya faruwa lokacin da Substrate a ƙasa da shawarar da aka ba da shawarar.
