Newcastle University Wetlab/3 September 2008

Wednesday 3rd September
Transformations


 * Today our aim was to transform our ligated plasmids into DH5-aplha E.coli competent cells.


 * For each transformation, we added 100µl of DH5-alpha E.coli competent cells to 12.5µl of the ligation mixtures from yesterday.


 * So in total we carried out 6 transformations, preparing them all on ice:

T1) Transforming pGFP-rrnB containing the 2.2kb fragment from pUC57-ncl08 into DH5-alpha

T2) Transforming pGFP-rrnB minus the 2.2kb fragment from pUC57-ncl08 into DH5-alpha

T3) Transforming pJWV021 containing the 2.2kb fragment from pUC57-ncl08 into DH5-alpha

T4) Transforming pJWV021 minus the 2.2kb fragment from pUC57-ncl08 into DH5-alpha

T5) Positive control: DH5-alpha cells plus pGFP-rrnB

T6) Negative control: DH5-alpha cells only


 * The transformations were then plated onto antibiotic agar:

T1 : +spec

T2 : +spec (small and large volume plated)

T3 : +amp and +kan

T4 : +amp +kan

T5 : +spec

T6 : +spec