Purification of DNA Restriction Digestion Reactions for Cloning

University of Birmingham
School of Biosciences
Postdoctoral Research Fellow

Overall

Quality of Results

Ease-of-Optimization

What do these ratings mean?
Write a Review

Company:

Promega

Product Name:

Wizard SV Gel and PCR Clean-Up System

Catalog Number:

A9282

Using Wizard SV Gel and PCR Clean-UP sytem, the plasmid (8 Kb) restricted with restriction enzymes in a 50 microliter reactions were purified to remove enzymes and buffer components in the restriction reaction. Reactions were eluted in 30 microliter nuclease free water and recovery was 80% (820 ng from 1 microgram restriction) with very good quality (260:280 of 1.88). Purified plasmid backbone was further used for ligating with DNA insert.

Experimental Design and Results Summary

Application

Purification of DNA restriction reactions for cloning

Starting Material

1 microgram of plasmid DNA restricted with two restriction enzymes in a total volume of 50 microliter per prep

Tips

None

Results Summary

The protocol very easy to follow. Just follow the instructions comes with kit's instruction manual. I started with 1 microgram of my plasmid DNA restricted with two restriction enzymes (plasmid size was 8 Kb). Before proceeding further for ligation of DNA insert, I needed to remove restriction enzymes and other salts in the restriction reaction. Using the kit, I have got purified plasmid backbone with good quality of 260:280 ratio of 1.88. Recovery was 80 % (recoverd 820 ng from 1 microgram plasmid restriction reaction).

DOI or PMID #

N/A

Image Gallery

Summary

The Good

Less hands-on time perform the purification (less than 15 mintues)

The Bad

None

The Bottom Line

Highly suggest this kit for purification of restriction reactions or PCR reactions for successful cloning method.

Join the discussion