DNA Miniprep
By Levi Clancy for Student Reader on
updated
- Genetic techniques
- 5'-Deletion Mutants
- Ames Test
- Cloning Vectors
- Conjugation
- DNA Fingerprinting
- DNA Miniprep
- Gel Shift Assay
- Gene Control in Development: Laboratory Techniques
- Gene Targeting
- Genetic Engineering
- Genetic screen
- In Vitro Nuclear Run-on Experiment
- Interrupted Mating Experiment
- Knockout mutation
- Linkage analysis
- Polymerase Chain Reaction
- Promoter (Transcriptional) (RNA) Fusion
- Reporter Gene
- Restriction Enzymes (Endonucleases )
- Sequence Alignment
- Shotgun sequencing
- Temperature Sensitive Mutant Experiment
- Transformation
- Transgenes
- Translational (Protein) Fusion
- Transposon Tagging
- cDNA Microarray
The goal of a DNA miniprep is to isolate only plasmid DNA from a prokaryote. The steps are to (a) lyse the cells (b) remove cellular membranes and waste (c) wash away undesired elements and (d) elute DNA. The elution is one of the most important steps. The negatively-charged DNA noncovalently binds to a filter, and the filter is washed. Theoretically, everything will flow through except for the DNA. After washing, the DNA is then removed from the filter (a process called elution). There are several buffers commonly used to elute and store the DNA. These buffers include EB, TE and dH2O.
There are many different kits used for minipreps. All these kits follow the same steps described above. A generalized procedure for a Qiagen miniprep is:
spin down cells
resuspend in buffer P1
add buffer P2; incubate
add buffer N3; centrifuge
run supernatant through column
wash column with buffer PE
elute DNA with buffer EB