"

From 2013.igem.org

Sample	Plasmid Volume(ul)	DNA concentration (ng/ul)
1.1	40	131.8
1.2	40	77.1
1.3	46	56.8
1.4	39	109
1.5	41	129.1
1.6	32	93.3

• measured DNA concentration using nanodrop

Restriction enzyme digest of the DNA samples

• • enzymes: XbaI, PstI
  • master mix:
    • Buffer - 14ul
    • H2O - 88.2ul
    • XbaI - 1.4ul
    • PstI - 1.4ul
    • Total volume - 105ul
• Each sample contained 5ul of DNA and 15ul of the master mix.
• The total amount of DNA in each sample was 200ng
• To calculate it we divided 200ng by the concentration of each sample
• The rest of the volume was made up with water to give a total of 5ul.
• The samples were then incubated in a water bath at 37C for an hour

Agorose Gel preparation

• • • 20ml of 5xTBE
    • 80ml of water
    • 0.8g of agarose
  • Boil for 3 minutes, 1 minute at a time.
  • Let it cool and add 5ul of ethidium bromide.
  • Pour and let it set for 30 mins.
  • Adding dye and marker
  • Used orange g loading dye.
    • 2ul to each sample
  • Used 1kb ladder, 5ul per 1 marker well.

• Run the gel
• Gel picture