Desalting Of Dagk Prep Of Samples For Faruv Cd Of Dagk Protocol

The key to far-UV CD is to have a clear solution which also does not have anything in it (besides protein) which absorbs in the 180-250nm range. Even if any such absorptive substance produces no CD signal, it can strongly absorb both components…

Glutaraldehyde Crosslinking Of Dagk Mutants Protocol

Purpose: Normal wild type DAGK will get covalently linked by glutaraldehyde to form primarily trimeric DAGK which can be observed in SDS-PAGE. This reflects the fact that DAGK functions as a homotrimer. Some DAGK mutants may not fold correctly…

Western Blot Detection Of Histagged Proteins Protocol

Use flat-bladed tweezers to handle the membrane. A small yellow tip box works well for this procedure. Gently agitate on platform shaker during each wash/incubation step. Leave for 10 minutes during the wash steps. Use about 30 mL of solution…

Bowie Procedure For Purifying Dagk Protocol

1. Lyse cells as described for the usual Sanders procedure. 2. To extract, add -octyl glucoside to 5% (5g/100 mL). 3. Tumble 30 min. (only) at 4C (do NOT use a stir bar, instead gently rotate a sealed container). The lysate cannot be frozen…

Regenerating Used Qiagen Ninta Resin Protocol

1. Pour used resin into filter funnel. May connect to vacuum line, but only if necessary. 25 2. Wash twice with 6M guanidine HCl + 2% formic acid. Stir well after EACH wash, and allow to drain completely between washes. 3.…

Ni2 Ninta Resin Incubation And Elution Protocol

1. Superflow. Nickel-Agarose resin should be equilibrated by rinsing with Buffer A. Use about 1.2 ml of packed resin for every gram of wet cells in the lysate. Pack resin into a column and rinse twice with 2 bed volumes of Buffer A. 2. Transfer…