Rnase Inhibitor
Murine RNase inhibitor is recombinant murine RNase inhibitor inoratidzwa nekucheneswa kubva kuE.coli.Inosungira kuRNase A, B kana C muchiyero che1: 1 kuburikidza nekubatana kusina-covalent, nokudaro kuvharidzira basa rema enzymes matatu uye kudzivirira RNA kubva pakuora.Nekudaro, Izvo hazvishande kurwisa RNase 1, RNase T1, S1 Nuclease, RNase H kana RNase kubva kuAspergillus.Murine RNase inhibitor yakaedzwa neRT-PCR, RT-qPCR uye IVT mRNA, uye yaienderana neakasiyana ekutengesa Reverse transcriptases, DNA polymerases uye RNA polymerases.
Inofananidzwa nevanhu RNase inhibitors, murine RNase inhibitor haina ma cysteine anonyanya kunetseka kune oxidation iyo inokonzera kusagadziriswa kweiyo inhibitor.Izvo zvichiita kuti zvigadzikane pakadzika kudzika kweDTT (pasi pe1 mM).Iyi ficha inoita kuti ive yakakodzera kushandiswa mukuita uko yakanyanya kuwanda kweDTT inopesana nemaitiro (semuenzaniso Real-time RT-PCR).
Application
Ichi chigadzirwa chinogona kushandiswa zvakanyanya mune chero kuyedza uko RNase kupindira kunogoneka kudzivirira kusvibiswa kweRNA, senge:
1.Synthesis yekutanga tambo ye cDNA, RT-PCR, RT-qPCR, nezvimwe.
2.Inodzivirira RNA kubva mukuora panguva ye in vitro transcription/ dudziro (semuenzaniso, in vitro viral replication system).
3.Kudzivirirwa kweRNase chiitiko panguva yekuparadzaniswa kweRNA uye kucheneswa.
Storage Conditions
Chigadzirwa chacho chinogona kuchengetwa pa -25～- 15 ℃, inoshanda kwemakore maviri.
Storage buffer
50 mM KCl, 20 mM HEPES-KOH (pH 7.6, 25 ℃), 8 mM DTT uye 50% glycerol.
Tsanangudzo yeyuniti
Nhamba yemurine RNase inhibitor inodiwa kuti inhibite basa re5ng ye ribonuclease A ne50% yakatsanangurwa seimwe unit (U).
Molecular uremu hweprotein
Huremu hwemurine RNase inhibitor ndeye 50 kDa.
Kudzora kwehutano
Exonuclease Chiitiko:
40 U yemurine RNase inhibitor ine 1 μg λ -Hind III digest DNA pa 37 ℃ kwemaawa 16 haibereki kusvibiswa sezvinotsanangurwa neagarose gel electrophoresis.
Endonuclease Chiitiko:
40 U ye murine RNase inhibitor ine 1μ g λ DNA pa 37 ℃ kwemaawa 16 haibudisi kuora sezvinotsanangurwa neagarose gel electrophoresis.
Nicking Chiitiko:
40U yemurine RNase inhibitor ine 1μ g pBR322 pa 37 ℃ kwemaawa 16 haibudisi kuora sezvinotsanangurwa neagarose gel electrophoresis.
RNase Chiitiko:
40U yemurine RNase inhibitor ine 1.6μ g MS2 RNA ye 4h pa 37 ℃ haibereki kuderedzwa sezvinotsanangurwa neagarose gel electrophoresis.
E.coli DNA:
40 U yemurine RNase inhibitor inoongororwa kuvapo kweE. coli genomic DNA uchishandisa TaqMan qPCR ine maprimers chaiwo eE. coli 16S rRNA locus.Iyo E. coli genomic DNA kusvibiswa ndeye ≤ 0.1 pg/40 U.
Notes
1.Violent oscillation kana kubvongodza kuchaita kuti enzyme inactivation.
2.The optimum tembiricha range yeiyi inhibitor yaive 25-55 ℃, uye Yakanga isina kuvharwa pa65℃ uye pamusoro.
3.Mabasa eRNase H, RNase 1 uye RNase T1 akanga asina kudziviswa nemurine RNase inhibitor.
4.Kuvharidzirwa kwebasa reRNase kwakawanikwa muhupamhi hwepH (pH 5-9 yose yakashanda), uye basa repamusoro rakaonekwa pa pH 7-8.
5.Sezvo ma ribonucleases achiwanzo chengetedza chiitiko pasi pemamiriro ekuita, kungwarira kunofanirwa kutorwa kudzivirira kumisa mamorekuru eRNase Inhibitor ayo akasanganiswa ne ribonuclease.Kuti udzivise kuburitswa kweribonuclease inoshanda, tembiricha dzinopfuura 50 ° C uye yakanyanya kuwanda yeurea kana mamwe madhinaturing agents anofanirwa kudzivirirwa.
