3.10 Nucleic Acid Probes
A similar replica plating procedure may be used if we have a radioactive
nucleic acid probe that will aneal with our new gene. The procedure is as
Figure 22. Screening with a radioactive probe
- Grow the cells on the master plate as before, but substitute a nitrocellulose
filter for the velvet cloth to blot the colonies.
- Press a nitrocellulose filter onto the colonies; some cells will adhere.
When the attached cells are treated with alkali, the cells will break open
and the DNA will attach to the nitrocellulose filter. A radioactive probe
derived from the new gene may then be incubated with the filters.
- Remove the filter with attached cells and treat with alkali.
- Incubate with a radioactive nucleic acid probe (RNA or DNA) under anealling
- Detect where the probe bound by autoradiography using X-Ray film.
The probe binds to those colonies that contain the new gene. We then
may go back to the master plate and select the desired colonies for further