Rnase A
I-Ribonuclease A (RNaseA) yi-polypeptide ene-stranded enye equkethe i-4 disulfide bond kunye ne-molecular weight malunga ne-13.7 kDa.I-RNase Ais i-endoribonuclease eyehlisa ngokukodwa i-RNA enemisonto enye kwi-C kunye neentsalela ze-U.Ngokukodwa, i-cleavage iyaqaphela ibhondi ye-phosphodiester eyenziwe yi-5'-ribose ye-nucleotide kunye neqela le-phosphate kwi-3'-ribose ye-nucleotide ekufutshane ye-pyrimidine, ukuze i-2,3'-Cyclic phosphates ifakwe kwi-hydrolyzed kwi-3 ehambelanayo. 'i-nucleoside phosphates (umz.,pG-pG-pC-pA-pG icandwa yiRNase A ukuvelisa i-pG-pG-pCp kunye ne-A-PG).I-RNase A yeyona nto isebenzayo ekucakeni i-RNA enomsonto omnye.Ugxininiso lokusebenza olucetyiswayo yi-1- 100 μ G/mL, iyahambelana neenkqubo ezahlukeneyo zokusabela.I-concentration yetyuwa ephantsi (i-0-100 mM NaCl) ingasetyenziselwa ukusika i-RNA enye, i-RNA ene-double-stranded, kunye ne-RNA ye-RNA eyenziwe yi-RNA-DNA hybridization.
Nangona kunjalo, kwi-concentration yetyuwa ephezulu (≥0.3 M), i-RNase A iqhekeza kuphela i-RNA enomtya omnye.
I-RNase A isetyenziswa ngokuqhelekileyo ukususa i-RNA ngexesha lokulungiswa kwe-plasmid DNA okanye i-genomic DNA.Ingaba okanye i-DNase iyasebenza ngexesha lenkqubo yokulungiselela inokuchaphazela ngokulula ukusabela.Indlela yendabuko yokubilisa kwindawo yokuhlambela amanzi ingasetyenziselwa ukungasebenzi komsebenzi we-DNase.Le mveliso ayinayo i-DNase kunye ne-protease, kwaye ayifuni unyango olushushu ngaphambi kokusetyenziswa.Ukongeza, le mveliso inokusetyenziswa kuvavanyo lwebhayoloji yemolekyuli njengohlalutyo lokhuseleko lwe-RNase kunye nohlalutyo lokulandelelana kwe-RNA.
Iimeko zokugcina
Imveliso kufuneka igcinwe kwi -25℃~-15℃ iminyaka emi-2.
Iinkcukacha
| Imbonakalo | Umgubo |
| Ubungakanani | 100mg/1g |
| Uhlobo lwemveliso | RNase A |
Imiyalelo
Le yenye yeendlela eziqhelekileyo zokulungiselela isisombululo se-RNase A.Isenokulungiswangezinye iindlela ngokweendlela zemveli elabhoratri okanye uncwadi lwereferensi (njengeukunyibilika ngokuthe ngqo kwi-10 mM Tris-HCl, pH 7.5 okanye isisombululo se-Tris-NaCl)
1. Sebenzisa i-10 mM i-acetate ye-sodium (pH 5.2) ukulungiselela i-10 mg / mL ye-RNase A isisombululo sokugcina.
2. Ukufudumala kwi-100 ℃ nge-15 min.
3. Ukuphola kwiqondo lobushushu begumbi, yongeza i-1/10 yevolumu ye-1 M Tris-HCl (pH 7.4), lungisa i-pH yayo kwi-7.4 (yeumzekelo, yongeza i-500 mL ye-10 mg / mL isisombululo sokugcina i-RNase 1 M Tris-HCl, pH7.4).
4. Ipakishwe ngaphantsi kwe -20℃ yokugcinwa emkhenkceni, enokuzinza ukuya kuthi ga kwiminyaka emi-2.
[Amanqaku]:
Xa ubilisa isisombululo se-RNaseA phantsi kweemeko ezingathathi hlangothi, imvula ye-RNase iya kwenza;Yibilise kwi-pH ephantsi, kwaye ukuba kukho imvula, inokubonwa, enokuthi ibangelwa ubukho bokungcola kweeprotheni.Ukuba intlenga ifunyenwe emva kokubiliswa, ukungcola kunokususwa ngesantya esiphezulu se-centrifugation (13000rpm), kwaye emva koko sipakishelwe indawo yokugcina umkhenkce.
Amanqaku
Nceda unxibe iPPE eyimfuneko, idyasi yelebhu kunye neeglavu ezinjalo, ukuqinisekisa impilo nokhuseleko lwakho.
