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Contents 1 Preparation for pH acclimatisation of Bacillus subtilis 168 2 Double digest to confirm filamentous cell part was cloned successfully 2.1 Aims 2.2 Materials and protocols 2.3 Results 2.4 Conclusion

Preparation for pH acclimatisation of Bacillus subtilis 168

We prepared 100mM of HEPES (it is a dibasic compound) at pH 7.0 and sent it for autoclaving.

Double digest to confirm filamentous cell part was cloned successfully

Aims

The aim of the experiment is to test if the filamentous cell part was cloned into pGFP-rrnB successfully.

Materials and protocols

1. Please refer to: Plasmid extraction.
2. Please refer to:Restriction digest. We used Nhe1 and Spe1 to remove the yneA from pGFP-rrnB.
3. Please refer to: Gel electrophoresis for running all the digested plasmid fragments.

Results

Figure 1: Gel electrophoresis result for restriction digest of pGFPrrnB and yneA with Nhe1 and Spe1.

• Lane 1: 1kb DNA ladder
• Lane 2: Tube 1
• Lane 3: Tube 2
• Lane 4: Tube 3
• Lane 5: Tube 4
• Lane 6: Tube 5
• Lane 7: Tube 6
• Lane 8: Tube 7
• Lane 9: Tube 8
• Lane 10: Tube 9
• Lane 11: Tube 10
• Lane 12: Tube 11
• Lane 13: Tube 12
• Lane 14: 1kb DNA ladder

Conclusion

The results show that the digest works, there is a band at approximately 541bp corresponding to yneA and a band at approximately 8.4kbp corresponding to pGFPrrnb in lanes 2, 3, 4, 6, 7, 8, 9, 11 and 12.