Bleomycin resistance gene
Resistance gene for algae expression. Bleomycin gives a resistance against zeocin, because ble binds to zeocin blocking its funtion.
Figure 1: Gel electrophoresis of mCherry+pSB1C3 construct
After ligating the sequence in the pJP22, we wanted to find the best way to transform C. reinhardtii, we tested Sapphire Blue, TAP medium and water as buffers during the electroporation. Then, cells were plated in Agar-TAP medium Petri dishes with different concentrations of Zeocin (an antibiotic from Bleomycin family) because the plasmid used had a sequence for Bleomycin resistance.
Figure 2: pJP22 mCherry transformants - Zeocin 5 ug/mL Figure 3: pJP22 mCherry transformants - Zeocin 10 ug/mL
Figure 4: pJP22 mCherry transformants - Z5 (Water Transformed) Figure 5: pJP22 mCherry transformants - Z10 (Water Transformed)
Figure 6: pJP22 mCherry transformants - Z5 (TAP Transformed) Figure 7: pJP22 mCherry transformants - Z10 (TAP Transformed)
pJP22 Figure 8: mCherry transformants - Z5 (Sapphire Transformed) Figure 9: pJP22 mCherry transformants - Z10 (Sapphire Transformed)
As expected, the number of colony-forming units is much higher in the media with less Zeomicin.
At the same time, the transformation of cells were not so good using Sapphire Buffer in comparison with Water and TAP medium.
Sequence and Features
- 10COMPATIBLE WITH RFC
- 12COMPATIBLE WITH RFC
- 21COMPATIBLE WITH RFC
- 23COMPATIBLE WITH RFC
- 25INCOMPATIBLE WITH RFCIllegal NgoMIV site found at 774
Illegal NgoMIV site found at 835
- 1000COMPATIBLE WITH RFC