DNA extraction procedure
Lysis1. V tubes obemom 1,5-2 mL place 300 uL krovi i add 900 uL RBC bufera.2. Vstryakhnut tubes on a vortex mixer.3. Incubate v techenie 5-10 minut at room temperature.4. Centrifuge v techenie 1 minuty pri maks. ob/min (ne menee 13 tys. ob/min dlya nastolnykh tsentrifug), dalee udalit supernatant.5. K poluchennomu osadku kletok add 300 uL liziruyushchego bufera i 20 uL Proteinazy K, shake vigorously on a vortex mixer until complete razrusheniya konglomerata kletok.6. Incubate v termostate pri 55-60℃ v techenie 15-25 minut, periodicheski vstryakhivaya.7. Perenesti tubes na led ili v kholodnyy shtativ i inkubirovat v techenie 1 minuty. V sluchae otsutstviya okhladitelnykh elementov inkubirovat at room temperature v techenie 5-10 minut until completely cooled smesi.Nucleic acid sorption and precipitation1. Add k smesi 50 uL osazhdayushchego bufera 1.2. Intensivno peremeshat tubes on a vortex mixer, sbrosit kapli s pomoshchyu brief centrifugation.3. Incubate v techenie 3-5 minut vo ldu ili v cooling rack. V sluchae otsutstviya okhladitelnykh elementov inkubirovat at room temperature v techenie 5-10 minut.4. Centrifuge tubes v techenie 5 minut pri 13 tys. ob/min.5. Akkuratno, ne zadevaya pellet, perenesti 300-330 uL supernatanta v novuyu tube obemom 1,5-2 mL.6. Add 330 uL osazhdayushchego bufera 2 i peremeshat s pomoshchyu pyatikratnogo perevorachivaniya tubes.7. Dlya uvelicheniya vykhoda DNA rekomenduetsya inkubirovat tubes v freezer pri -20℃ v techenie 25 minut.8. Centrifuge tubes v techenie 5 minut pri 13 tys. ob/min, posle chego udalit supernatant.Nucleic acid washing1. Add v tubes 700 uL promyvochnogo bufera 1.2. Intensivno peremeshat tubes on a vortex mixer.3. Centrifuge tubes v techenie 5 minut pri 13 tys. ob/min, posle chego udalit supernatant.4. Add v tubes 700 uL promyvochnogo bufera 2.5. Repeat punkty 2,3.6. Vysushit samples s otkrytymi kryshkami v termostate pri 42℃ ili v tsentrifuge-kontsentratore pri 30℃ until complete ispareniya promyvochnogo rastvora.Nucleic acid elution1. Add v tubes 50-100 uL elyuiruyushchego bufera.2. Incubate v termostate pri 60℃ v techenie 5 minut, periodicheski peremeshivaya.Poluchennye rastvory nucleic acids can be stored do 7 dney pri temperature ot +2℃ do +4℃ i do dvukh let pri temperature -20℃. Po absorbance ratio na A260/A280 purity of the resulting solution DNA sootvetstvuyut ≥1,7.
return to product catalog