Proteinase K NGS (pulveris)
Cat No: HC4507A
NGS Protease K est protease seria stabilis cum actione enzyme alta et specificitate substrata late. Enzyma potiore dissolvit vincula et vincula peptide cum C-terminalibus acida hydrophobica amino, acida sulphurea continente amino acida et amino aromatica.Ita saepe servom in breves peptides detrahendum est.NGS Protease K est typicam serinam cum Aspis39-His69-Ser224trias catalytica, quae unica est ad serinam proteastionem, et centrum catalyticum cingitur stuppa Ca.2+sites ligandi ad stabilizationem pertinentes, altam enzyme navitatem sub condicionibus latius sustinendis.
Specification
| Aspectus | Alba ad extemporalitatem amorpho pulveris albi, lyophilized |
| Imprimis actio | ≥40U/mg solida |
| DNase | Nihil detectum |
| RNase | Nihil detectum |
| Bioburden | ≤50CFU/g solidum |
| Residuum acidum nucleicum | <5pg/mg solidum |
Properties
| Source | Tritirachium album |
| EC numerus | 3.4.21.64(Recombinant from Tritirachium album) |
| M. pondus | 29kDa (SDS-PAGE) |
| Isoelectric punctum | 7.81 Fig.1 |
| optimum pH | 7.0-12.0 (Omnes alta actione praestare) Fig.2 |
| Optimum temperatus | 65℃ Fig.3 |
| pH Stabilitas | pH 4.5-12.5 (25℃,16h) Fig |
| Scelerisque status | Infra 50℃ (pH 8.0, 30min) Fig |
| At stabilitas | Condita in 25℃ pro 12months Fig.6 |
| Activators | SDS, urea |
| Inhibitors | Diisopropyl fluorophosphate;benzylsulfonyl fluoride |
Repono Conditions
Condunt lyophilized pulverem -25~-15 diu a luce abductum;Post dissolutionem aliquot in opportunitate voluminis ad breve tempus repositionis 2-8℃ a luce vel diuturno tempore ad -25~-15 a luce.
Cautiones
Gerunt chirothecas tutelares et goggles cum utendo vel appendendo, et post usum bene ventilatas.Productum hoc facere potest cutis reactionem allergic et gravem oculi irritationem.Si haustus, allergiam vel asthma signa vel dyspnoeam causare potest.Sit irritatio respiratorii causa.
Unitas definition
Una unitas NGS Proteasionis K definitur quantitate enzyme ad hydrolyandam casein in 1 μmol L-tyrosine sub norma determinationis conditionibus.
Reagentia praeparatio
| Reagent | Manufacturer | Catalogue |
| Casein technicade lacte bubulo | Sigma Aldrich | C7078 |
| NaOH | Sinopharm ChemicalReagent Co., Ltd. | 10019762 |
| NaH2PO4·2H2O | Sinopharm ChemicalReagent Co., Ltd. | 20040718 |
| Na2HPO4 | Sinopharm ChemicalReagent Co., Ltd. | 20040618 |
| Acidum trichloroaceticum | Sinopharm ChemicalReagent Co., Ltd. | 80132618 |
| Natrium acetate | Sinopharm ChemicalReagent Co., Ltd. | 10018818 |
| Acidum acetic | Sinopharm ChemicalReagent Co., Ltd. | 10000218 |
| HCl | Sinopharm ChemicalReagent Co., Ltd. | 10011018 |
| Sodium carbonas | Sinopharm ChemicalReagent Co., Ltd. | 10019260 |
| Foline-phenol | Sangon Biotech (Shanghai)Co., Ltd. | A500467-0100 |
| Dominus tyrosine | Sigma | 93829 |
Reagent I:
Substratum: 1% Casein ex solutione lac bovis: solve 1g caseinum lac bovinum in 50ml solutionis phosphatarum sodium 0.1M, pH 8.0, caloris in aqua balnei ad 65-70 °C pro 15 min, move et dissolve, cum aqua refrigerandi, adaequatum. sodium hydroxidum ad pH 8.0, et dilutum ad 100ml.
Reagent II:
Solutio TCA: acidum trichloroaceticum 0.1M,0.2M natrium acetatis et 0.3M aceticum aceticum (pondere 1.64g acidum trichloroaceticum + 1.64g acidum aceticum + 1.724mL Acidum aceticum successive, adde aquam deionizatam 50mL, compone cum HCl ad pH 4.03, ac dilue in 100ml).
Reagent III:
0.4m Solutio aphronitri aphronitri (ponde 4.24g aphronitri anhydrous aphronitri et solve in aqua 100mL)
Reagens IV:
Folin phenol reagens: dilutum 5 temporibus cum aqua deionized.
Reagens V:
Enzyme diluens: 0.1 M solutio phosphatae sodium, pH 8.0.
Reagens VI:
Solutio vexillum L-tyrosinum: 0, 0.005, 0.025, 0.05, 0.075, 0.1, 0,25 umol/ml L-tyrosinum dissolutum cum 0.2M HCl.
De modo procedendi
1. In UV-Vis spectrophotometrum converte et mensurae photometricae selectae.
2. 660nm constitue = the wavelength.
III. Convertere in aquam balneum, tempera temperatura 37℃, curet temperatura immutata 3-5 min.
4. Preheat 0.5mL distent in tubo 2mL centrifugii ad 37℃ aquae lavacrum pro 10min.
5. Extract 0.5mL solutio enzyme diluta in tubum centrifugium preheatum per 10 minutas.Set enzyme diluent ut blank group.
6. Adde 1.0 mL TCA statim post reactionem reagens.Misce bene et incubare in aqua balnei pro XXX minutis.
7. solutio reactionis centrifugae.
8. Adde sequentia membra ordine determinato.
| Reagent | Magnitudo |
| Supernatant | 0.5 mL |
| 0.4M Sodium carbonas | 2.5 mL |
| Folin phenol reagens | 0.5 mL |
9. Misce bene antequam incubans in aqua balnei 37℃ pro 30 min.
10. OD660determinari ad OD1;blank control group: Enzyme diluent enzyme solution ad determinare OD reponere adhibetur660ut OD2, OD=OD1-OD2.
11. L-tyrosinum vexillum curvae: 0.5mL diversae solutionis concentrationis L-tyrosinae, 2.5mL 0.4M Sodium carbonas, 0.5mL Folin phenol reagens in 5mL tubi centrifugii, incubare 37℃ pro 30min, deprehendere pro OD.660ob diversam intentionem L-tyrosinam, tum curvae regulae Y=kX+b consecuta, ubi Y est L-tyrosina concentratio, X est OD.600.
Calculus
2: Totalis voluminis solutionis reactionis (ml)
0.5: Volume enzyme solutionis (mL)
0,5: Reactio liquida volumen in determinatione chromogenic (mL)
X: Reactio tempus (min)
Df: Dilutio multiplex
C: Enzyme concentration (mg/mL)
Figurae
Fig.1 DNA residua
| Sample | Ave C4 | Acidum nucleicum Recuperatio (pg/mg) | Recuperatio (%) | Totalis Nucleic Acidum ( pg/mg) |
| PRK | 24.66 | 2.23 | 83% | 2.687 |
| PRK+STD2 | 18.723 | 126.728 | - | - |
| STD1 | 12.955 |
- |
- |
- |
| STD2 | 16 | |||
| STD3 | 19.125 | |||
| STD4 | 23.135 | |||
| STD5 | 26.625 | |||
| RNA-Free H2O | indeterminatum | - | - | - |
Fig.2 Optimum pH
Fig.3 Optimum temperatus
Fig.4 pH Stabilitas
Fig.5 Scelerisque stabilitas
Fig.6 at stabilitas 25℃
