Reagent kit for plasmid DNA extraction from bacterial culture "Plasmidex Midi" Raissol

V nabor vklyucheny vse neobkhodimye reagenty dlya extraction plazmidnoy DNA:
1. Resuspendiruyushchiy bufer - dlya razrusheniya konglomerata kletok;
2. Lysis buffer - na osnove chaotropic agents i detergentov dlya optimal lysis i denaturatsii genomnoy DNA;
3. Neytralizuyushchiy bufer - dlya stabilizatsii pH rastvora;
4. Promyvochnyy rastvor - dlya promyvki nucleic acids ot cell metabolites and salts;
5. Elution buffer - dlya elyutsii i khraneniya nucleic acids, pH=8,9;
6. RNase A - dlya udaleniya RNA;
7. Spin-kolonki - dlya sorbtsii NA.

Download the short instruction "Plasmidex Midi" (pdf)

Download the full instruction "Plasmidex Midi" (pdf)

Podgotovka rastvorov
Add ves obem RNAazy A v resuspendiruyushchiy bufer.
Optsionalno: dlya povysheniya sroka khraneniya rastvorov, RNase A mozhet dobavlyatsya v sample individualno v obeme 5 uL posle vneseniya resuspendiruyushchego bufera.

Provedenie protsedury extraction plazmidnoy DNA

Lysis
1. V tubes obemom 15 mL vnesti 10 mL nochnoy bakterialnoy kletochnoy kultury i tsentrifugirovat v techenie 6 minut pri 3-5 tys. ob/min. Remove the supernatant. Pri neobkhodimosti povysheniya kontsentratsii tselevogo produkta povtorit operatsiyu, dobavlyaya k osadku kletok svezhuyu bakterialnuyu kulturu s obshchim obemom kultury do 15 mL.
2. Add k osadku 1 mL resuspendiruyushchego bufera, peremeshat on a vortex mixer ili pipetirovaniem until complete razrusheniya konglomerata kletok.
3. Vnesti 1 mL liziruyushchego bufera. Mix s pomoshchyu plavnogo perevorachivaniya tubes ili plavnym pipetirovaniem do prozrachnosti rastvora. Incubate v techenie 2-3 minut at room temperature. Ne vortex mixerirovat.
4. Add 1,5 mL neytralizuyushchego bufera. Mix s pomoshchyu plavnogo perevorachivaniya tubes ili plavnym pipetirovaniem do obrazovaniya tvorozhistoy vzvesi. Ne vortex mixerirovat. Incubate at room temperature v techenie 3-5 minut.
Nucleic acid sorption and precipitation
1. Centrifuge tubes v techenie 30 minut na maksimalnoy skorosti.
2. Ne zadevaya pellet, perenesti 3 mL supernatanta v podgotovlennuyu spin-kolonku.
3. Centrifuge spin-kolonki v techenie 1 minuty pri maks. ob/min. Udalit filtrat iz sobiratelnykh probirok.
Nucleic acid washing
1. Add 3 mL promyvochnogo rastvora i tsentrifugirovat samples v techenie 1 minuty pri maks. ob/min. Udalit filtrat iz sobiratelnykh probirok.
2. Repeat punkt 1.
3. Vernut spin-kolonki v sobiratelnye tubes. Centrifuge tubes v techenie 1 minuty pri maks. ob/min dlya udaleniya ostatkov promyvochnogo rastvora.
Nucleic acid elution
1. Utilizirovat sobiratelnye tubes, perenesti spin-kolonki v novye tubes obemom 15 mL.
2. Nanesti v tsentr membrany spin-kolonok 250 uL elyuiruyushchego bufera, inkubirovat v techenie 2-5 minut at room temperature.
3. Centrifuge tubes v techenie 1 minuty pri maks. ob/min.
4. Utilizirovat spin-kolonki.

Buffers nabora "Plasmidex Midi" can be stored at room temperature for 12 months from the manufacturer's release date. If a precipitate is present pri minimalnykh temperaturakh khraneniya warm the solutions to room temperature.
Enzyme RNase A khranitsya pri -20℃.
Posle dobavleniya RNAazy A v resuspendiruyushchiy bufer poluchennyy rastvor sleduet khranit pri +4℃ ne bolee 6 mesyatsev.
Poluchennye rastvory plazmidnoy DNA can be stored do 7 dney pri temperature ot +2℃ do +4℃ i do dvukh let pri temperature -20℃.

Optsionalno: dlya povysheniya sroka khraneniya rastvorov, RNase A mozhet dobavlyatsya v sample individualno v obeme 5 uL posle vneseniya resuspendiruyushchego bufera.