IGEM:Peking/2007/Switch-Notebook/2007-8-9

miniprep of T1TE-plx010 and GFP-plx003

digestion test:

T1TE-plx010 10uL

ScaI/XhoI 0.5uL each

H buffer 2uL

add water to a total 20uL

GFP-plx003 10uL

ScaI/XhoI 0.5uL each

H buffer 2uL

add water to a total 20uL

result: GFP does not digest well.

Colony PCR

GFP-plx003: primer of testF and GFP-R

T1TE-GFP-Plx007: primer of T1TE(R) and T1TE(lacZ)-F

T1TE-T1T2-pcc010: primer of pkanF and T1T2-R

PCR double promoters

plac-T, prec-T, psul-T

SS1-T, SS2-T, SS3-T, SD1-T,SD2-T,SD3-T

using gel rescue to get many double promoter fragments for next step

GFP-plx007 plasmid double digestion

plasmid 2uL

XbaI/XhoI 2uL each

buffer M 10uL

H2O 84uL

total 100uL system

fragment digestion

fragment from PCR rescue 16uL

H buffer 2uL

XhoI/SpeI 2uL each

ddH2O 70uL

total 100uL system