Please use The PureLink™ HiPure Plasmid DNA Miniprep Kit.

or try the following protocol:

1.   From a single white colony (truly white) on plates containing Blue-gal and IPTG, inoculate a 4 ml LB medium supplemented with appropriate antibiotics. Grow at 37°C to stationary phase (up to 24 hours) with shaking

2.   Transfer 1.5 ml of culture to a 1.5 ml microcentrifuge tube and centrifuge at 14,000xg for 1 min.

3.   Remove the supernatant and resuspend each pellet in 0.3 ml of solution 1 (15 mM Tris HCL pH 8.0, 10 mM EDTA, 100 µg/ml RNase A). Add 0.3 ml of 0.2 M NaOH, 1% SDS solution and gently mix. Incubate at room temperature for 5 mins.

4.   Slowly add 0.3 ml of 3 M potassium acetate pH 5.5, mixing gently during addition. A thick white precipitate of protein and E. coli genomic DNA will form. Place the sample on ice for 5 to 10 min.

5.   Centrifuge for 10 mins at 14,000xg. During the centrifugation, label another microcentrifuge tube and add 0.8 ml isopropanol to it. Gently transfer the supernatant to the tube containing isopropanol. Avoid any white precipitate material. Mix by gently inverting tube a few times and place on ice for 5 to 10 mins or store at -20°C overnight. Centrifuge the sample for 15 mins at 14,000 x g at room temperature.

6.      Remove the supernatant and add 0.5 ml 70% ethanol to each tube. Centrifuge for 5 mins at 14,000xg at room temperature.

7.      Remove as much of the supernatant as possible.

8.      Air dry the pellet briefly, 5 to 10 mins, at room temperature and dissolve the DNA in 40 µl sterile water. Store at -20°C.

Hi Alexandra

As mentioned above and recommended by the manufacturer , Purelink Hipure plasmid DNA minprep kit , Although the company say that it is the kit of choice but in my experience the BACMID DNA yield was too low and transfection of such DNA didn't work well.

I also used "QIAGEN Plasmid Maxi Kit " , it gave me a very good results after transfection.

There is protocol which do not need any kit, and very reliable according to our usage. It is published in

https://www.researchgate.net/publication/282755508_Recombinant_Expression_and_Reconstitution_of_MultiProtein_Complexes_by_the_USER_Cloning_Method_in_the_Insect_Cell-Baculovirus_Expression_System

2.4.2. Amplification of recombinant MultiBac bacmid DNA (step 3continued)In 50 ml falcon tubes, 10 ml of LB broth containing appropriateantibiotics (Table 6) is inoculated with 3–6 selected colonies. Cul-ture the inoculum at 37°C over night by shaking at 220 rpm. The cells are pelleted and resuspended in 0.5 ml suspension buffer(50 mM Tris–HCl pH 8.0, 10 mM EDTA, 100lg/ml RNaseA). Nextadd 0.5 ml of lysis buffer (200 mM NaOH, 1% SDS) to lyse the cellsthrough gentle mixing. Thereafter, handle with care to avoid shear-ing the bacmid DNA. Add 0.7 ml of neutralizing buffer (4.2 Mguanidinium chloride, 0.9 M potassium acetate, pH 4.8). Spin downthe lysate at 13,000 rpm for 10 min in a bench top centrifuge.Carefully pipette the supernatant into a new tube containing 1 vol-ume of isopropanol. Mix gently and precipitate the MultiBac bac-mid DNA by centrifugation for 5 min at high speed. Discard thesupernatant and spin down the DNA pellet briefly, the last traceof solution is aspirated by pipetting. The still wet bacmid DNApellet is gently resuspended with 100ul of sterile distilled water.

Article Recombinant Expression and Reconstitution of MultiProtein Co...

please send your e-mail address to me so I can forward my protocol for isolation.