Revision as of 17:23, 25 October 2010

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Problem

Genomic DNA extraction from Bacillus subtilis strain ATCC 6633

Aim

The aim of this experiment was to extract genomic DNA from Bacillus subtilis strain ATCC 6633. This strain produces the subtilin genes, required for our Subtilin Production and Immunity BioBricks as well as the Quorum sensor biobrick designed by Team Newcastle 2008.

Materials and methods

ara forward and reverse primers
sac forward and reverse primers
Pipettes
Universal tubes
Eppendorf tubes
Centrifuge
PCR (see PCR protocol from Lab training)
Gel electrophoresis (see Gel electrophoresis from Lab training)

ara and sac are two genes that exist in the ATCC 6633 strain. ara and sac forward and reverse primers are two tests, which will be used in PCR. At the end, Gel Electrophersis can be used to distinguish the bands. If the bands from either of ara or sac show up at the expected bps against the DNA ladder, this proves that the genomic DNA is sucessfully extracted.

Protocol

Please refer to: DNA extraction of Bacillus subtilis for materials required and protocol.

Results

Results were as expected see: https://2010.igem.org/Team:Newcastle/7_July_2010

Conclusion

See: https://2010.igem.org/Team:Newcastle/7_July_2010

Research

Today we also continued with our research on filamentous cells which will act as reinforcing fibres in the crack.

Dajkovic A., Mukherjee A. and Lutkenhaus J. (2008). "Investigation of Regulation of FtsZ Assembly by SulA and Development of a model for FtsZ Polymerization". Journal of Bacteriology". 190 (7). 2513- 2526.
Kawai Y. and Ogasawara N. (2006). "Bacillus subtilis ErzA and FtsL synergistically regulate FtsZ ring dynamics during cell division". Microbiology". 152. 1129- 1141.

@@ Line 27: / Line 27: @@
 See: https://2010.igem.org/Team:Newcastle/7_July_2010
-=LacI BioBrick Construction=
+=Research=
+Today we also continued with our research on filamentous cells which will act as reinforcing fibres in the crack.
-==Aims==
+#Dajkovic A., Mukherjee A. and Lutkenhaus J. (2008). "Investigation of Regulation of FtsZ Assembly by SulA and Development of a model for FtsZ Polymerization". ''Journal of Bacteriology". 190 (7). 2513- 2526.
-Use PCR to extract ''lacI'' (promoter, ribosome-binding site (RBS) & coding sequence (CDS)) from plasmid pMutin4 and ligate into vector pSB1AT3 in front of red fluorescent protein (''rfp'').
+#Kawai Y. and Ogasawara N. (2006). "''Bacillus subtilis'' ErzA and FtsL synergistically regulate FtsZ ring dynamics during cell division". ''Microbiology". 152. 1129- 1141.
-==Materials==
-* [[Team:Newcastle/5_July_2010#LacI_BioBrick_Construction|Overnight culture]].
-* Restriction enzymes EcoR1 and Xba1.
-* PCR primers designed for ''lacI'' extraction.
-==Protocol==
-* [[Team:Newcastle/Qiagen_Minipreps|Plasmid miniprep]] from overnight cultures.
-* [[Team:Newcastle/Restriction_digests|Restriction digest]] of pSB1AT3 with EcoR1 and Xba1.
-* Amplify ''lacI'' fragment by [[Team:Newcastle/PCR|PCR]] from pMutin4.
-* [[Team:Newcastle/Restriction_digests|Restriction digest]] of amplified fragment of pMutin4 with EcoR1 and Spe1.
-==Inference==
-* The plasmids are extracted from the transformed ''E. coli'' and digested so that the amplified ''lacI'' fragment can be inserted. Set up PCR to amplify the ''lacI'' promoter and coding sequence from pMutin4.
 {{Team:Newcastle/footer}}

Team:Newcastle/6 July 2010

From 2010.igem.org