EDVOTEK® BIOTECHNOLOGY BOOTCAMP
Bacterial Transformation and Transformation Efficiency
SOURCE PLATES: THE FOUNDATION TO A SUCCESSFUL TRANSFORMATION!
Over the years, we have improved our protocols to provide consistent results. We h ave found that a well-streaked source plate is one of the most important parts in guaranteeing a successful transformation!
What is a Source Plate?
A source plate is a standard LB Agar plate that is used to grow the initial colonies of bacteria that will be used for the transformation process. Source plates allow scientists to observe the growth of the bacteria to make sure it is grow-ing well and there is no contamination. Because of the way source plates are streaked, students are able to select well-isolated colonies to get the proper amount of bacteria for their transformation! We have found that when the source plates are older than 24 hours, and when clumps of bacteria are used rather than individual colonies, transformation is far less likely to succeed. Always pick the well-isolated colonies on your plate that are 1-1.5 mm in diameter.
Source Plates Should Be:
• Prepared the day before the lab and incubated overnight at 37ºC. • Used as soon as possible after the incubation period (18-22 hours) has finished. • Streaked properly to ensure the growth of well-isolated colonies (see figure, below).
FIGURE 1
Cover.
90°
90°
90°
Invert & incubate.
Add BactoBead™.
Rehydrate.
Streak 1st quadrant.
Rotate. Streak 2nd quadrant.
Rotate. Streak 3rd quadrant.
Rotate. Isolate colonies.
Streaking Source Plates:
1. Add a fresh BactoBead™ to an LB Agar plate. 2. Rehydrate the BactoBead™ with 10 µL of recovery broth. 3. Start a dense patch of bacteria in the first quadrant by spreading the BactoBead™ back in forth. 4. Rotate the plate 90º and drag the loop through the bacteria in the first quadrant. Streak back and forth making sure not to dip back into the previous quadrant. 5. Repeat this on the 3rd quadrant.
6. Drag the loop across the rest of the empty space on the agar to isolate colonies. 7. Cover and invert the plates, then place them in a 37ºC incubator overnight.
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