Miniprep completion, Gel #3 & sequencing

Contributors: Esther, Chloe

Miniprep — completed

Standard alkaline lysis/spin column protocol: resuspension, lysis, neutralisation, spin column wash ×2, elution.

Gel #3 — restriction digest verification

Digested samples in thermoshaker at 37 °C for ~2 hours. Lanes run: ladder, uncut / NcoI / EcoRI / NcoI+EcoRI digests for both CxnA-1 and CxnA-Hist (20 µl per lane, 10 µl ladder). Run at 100 V for 45 min; dye line ~75–80% down gel.

(Gel #3 produced interpretable bands — improvement over Gels #1 and #2.)

Nanodrop

Sequencing submission

Plasmid DNA prepared and submitted for Sanger sequencing. Dilution: C₁V₁ = C₂V₂ — 11 µl miniprep + 34 µl nuclease-free water = 45 µl total.