ESO wikiadmin: Created page with "{| class="wikitable" |+ !Materials |- |Restriction enzyme stock (–20°C) |- |Appropriate 10x enzyme buffer (–20°C) |- |Sterile dd H₂O |- |Depending on the digestion '''(optional)''': '''''BSA (bovine serum albumin)''''': To ensure 100% activity of the enzymes, 100 µg/ml BSA should be added (stored at –20°C) |} == Procedure for analysis purposes == * 300 ng of DNA samples (e.g. control digest of plasmid minipreps) * 1 µl appropriate enzyme buffer (1..."

2025-05-21T13:14:00Z

Created page with "{| class="wikitable" |+ !Materials |- |Restriction enzyme stock (–20°C) |- |Appropriate 10x enzyme buffer (–20°C) |- |Sterile dd H2O |- |Depending on the digestion '''(optional)''': '''''BSA (bovine serum albumin)''''': To ensure 100% activity of the enzymes, 100 µg/ml BSA should be added (stored at –20°C) |} == Procedure for analysis purposes == * 300 ng of DNA samples (e.g. control digest of plasmid minipreps) * 1 µl appropriate enzyme buffer (1..."

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{| class="wikitable"
|+
!Materials
|-
|Restriction enzyme stock (–20°C)
|-
|Appropriate 10x enzyme buffer (–20°C)
|-
|Sterile dd H2O
|-
|Depending on the digestion '''(optional)''':
'''''BSA (bovine serum albumin)''''': To ensure 100% activity of the enzymes, 100 µg/ml BSA should be added (stored at –20°C)
|}

== Procedure for analysis purposes ==

* 300 ng of DNA samples (e.g. control digest of plasmid minipreps)
* 1 µl appropriate enzyme buffer (10x)
* 0.1 µl restriction enzyme 1 (1 U)
* 0.1 µl restriction enzyme 2 (1 U)
* '''opt.''' 1 µl BSA (10 mg/ml stock solution)
* Fill up to 10 µl with ddH2O
* Incubation for 3 h at recomended temperature

== Procedure for preparative purposes (Overnight digestion) ==

* 2 – 4 µg of DNA sample (e.g. plasmid for preparative procedures)
* 2 µl appropriate buffer (10x)
* 0.2 µl restriction enzyme 1
* 0.2 µl restriction enzyme 2
* '''opt.''' 2 µl BSA (10mg/ml stock solution)
* Fill up to 20 µl with dd H2O
* ON incubation at recommended temperature

Restriction digest of plasmid DNA - Revision history