I-Rnase Inhibitor (iGlycerol mahhala)
I-Murine RNase inhibitor iyi-recombinant murine RNase inhibitor evezwe futhi yahlanzwa ku-E.coli.Ibophezela ku-RNase A, B noma C ngesilinganiso esingu-1:1 ngokusebenzisa i-non-covalent bonding, ngaleyo ndlela ivimbele umsebenzi wama-enzyme amathathu futhi ivikele i-RNA ekonakalisweni.Nokho, Ayisebenzi uma iqhathaniswa ne-RNase 1, RNase T1, S1 Nuclease, RNase H noma i-RNase evela ku-Aspergillus.I-Murine RNase inhibitor yahlolwa yi-RT-PCR, RT-qPCR kanye ne-IVT mRNA, futhi yayihambisana nezinhlobonhlobo ze-Reverse transcriptases zezentengiselwano, ama-DNA polymerases nama-polymerase e-RNA.
Uma kuqhathaniswa nama-RNase inhibitors abantu, i-murine RNase inhibitor ayinawo ama-cysteines amabili azwela kakhulu ku-oxidation ebangela ukungasebenzi kwe-inhibitor.Lokho kuyenza izinze ekugxilweni okuphansi kwe-DTT (ngaphansi kuka-1 mM).Lesi sici siyenza ifanelekele ukusetshenziswa ekuphenduleni lapho ukugxila okuphezulu kwe-DTT kuphambene nokusabela (isb. I-Real-time RT-PCR).
Aisicelo
Lo mkhiqizo ungasetshenziswa kakhulu kunoma yikuphi ukuhlola lapho ukuphazamiseka kwe-RNase kungenzeka khona ukuze kugwenywe ukucekelwa phansi kwe-RNA, okufana nalokhu:
1.Ukuhlanganiswa kwe-cDNA yomucu wokuqala, i-RT-PCR, i-RT-qPCR, njll.;
2.Vikela i-RNA ekonakalisweni kwe-in-vitro transcription/translation (isb. ukuphindaphinda kwegciwane ku-vitro);
3.Ukuvinjelwa komsebenzi we-RNase ngesikhathi sokuhlukaniswa kwe-RNA nokuhlanzwa.
Izimo Zokugcina
Gcina ku -25～-15℃;
Imijikelezo yokuqhwaza ≤ izikhathi ezi-5;
Ivumeleke unyaka ongu-1.
Incazelo yeyunithi
Iyunithi eyodwa ichazwa njengenani le-RNase Inhibitor elidingekayo ukuze kuvinjwe umsebenzi we-5 ng we-RNase A ngo-50%.
Isisindo samangqamuzana
I-RNase Inhibitor (i-Glycerol-free) iyiphrotheni engama-50 kDa.
Ikhwalithi yokulawula
I-Exonuclease Umsebenzi:
Ukufakwa kwe-40 U ye-enzyme ene-1 μg λ-Hind III yokugaya i-DNA amahora angu-16 ku-37°C kubangele kungabikho ukuwohloka okubonakalayo kwe-DNA njengoba kunqunywa ijeli electrophoresis.
Umsebenzi we-Endonuclease:
Ukufakwa kwe-40 U ye-enzyme ne-1 μg λ DNA amahora angu-16 ku-37°C kubangele kungabikho ukuwohloka okubonakalayo kwe-DNA njengoba kunqunywe ijeli electrophoresis.
Ukudlala Umsebenzi:
Ukufakwa kwe-40 U ye-enzyme ne-1 μg pBR322 amahora angu-16 ku-37℃ kubangele kungabikho ukuwohloka okubonakalayo kwe-DNA njengoba kunqunywe i-gel electrophoresis.
RNase Umsebenzi:
Ukufakwa kwe-40 U ye-enzyme ne-1.6 μg MS2 RNA amahora angu-4 ku-37℃ kubangele kungabikho ukuwohloka okubonakalayo kwe-RNA njengoba kunqunywe i-gel electrophoresis.
E.coli DNA:
I-40 U ye-Enzyme itholwa yi-TaqMan qPCR.I-E.coli DNA ingu-≤ 0. 1pg/40U.
Nothis
1.Unganyakazi noma unyakaze ngamandla ukuze uvimbele ukungasebenzi kwe-enzyme.
I-2.RNase inhibitor iyasebenza emazingeni okushisa asukela ku-25℃ kuya ku-55℃ futhi ayisebenzi kokuthi ≥65℃.
3.Umsebenzi we-RNase H, i-RNase 1, ne-RNase T1 awuvinjiwe.
4.Ibanga le-pH lokuvimbela umsebenzi we-RNase libanzi (lisebenza ku-pH 5-9), libonisa umsebenzi omkhulu ku-pH 7-8.
