Part:BBa_K2136014

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.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 774
  Illegal NgoMIV site found at 835
  
• 1000
  COMPATIBLE WITH RFC[1000]