Streptococcus suis Transformation: Difference between revisions
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#Add 0.5pMol of DNA (1.4ug for pACYC194; see https://www.promega.com/resources/tools/biomath/) | #Add 0.5pMol of DNA (1.4ug for pACYC194; see https://www.promega.com/resources/tools/biomath/) | ||
#Vortex; place in 37 degree incubator for 2 hours. | #Vortex; place in 37 degree incubator for 2 hours. | ||
#Plate onto appropriate antibiotic plates. | #Plate onto appropriate antibiotic Tryptic Soy plates. | ||
Latest revision as of 12:57, 16 December 2022
Day 1:
- Plate strains from freezer onto Tryptic Soy Plates for single colonies.
Day 2:
- Put 3ml of Todd-Hewitt + 0.5% Yeast Extract into sterile 16mm test tubes. Add a single colony into the tube; grow overnight at 37 degrees in the plate incubator (so, without shaking).
Day 3:
- Put 5ml of Todd-Hewitt + 0.5% Yeast Extract into sterile 16mm test tubes to pre-warm the media for 1 hour.
- Add 125 ul of the overnight growth (a 1:40 ratio).
- Allow to grow to an Absorbance of 0.035 - 0.058 (one hour, generally).
- Put 100ul of this culture into a micro-centrifuge tube.
- Add in the correct XIP peptide from the stock to create a final concentration of 250uM. (normally, 7-10ul of stock peptide)
- Add 0.5pMol of DNA (1.4ug for pACYC194; see https://www.promega.com/resources/tools/biomath/)
- Vortex; place in 37 degree incubator for 2 hours.
- Plate onto appropriate antibiotic Tryptic Soy plates.