Team:Newcastle/3 August 2010


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Plasmid Miniprep Experiment


The aim of this experiment is to extract plasmid DNA pSB1C3 and pSB1AK3 with BBa_B0014 from E. coli DH5α cells using the Qiagen miniprep kit and analysing with the Nanodrop machine.

Materials and Protocol

Please refer to: Minipreps for Qiagen miniprep protocol, Nanodrop Spectrophotometer for nanodrop protocol and Restriction digests for restriction digestion protocol. NOTE: 10 µl of RNAse A have been added into the current P1 buffer from Qiagen.



Figure 1: Gel electrophoresis of the PCR products

  • Lane 1: 1kb DNA ladder
  • Lane 2: Extraction of pSB1C3 plasmid (No. 1)
  • Lane 3: Extraction of pSB1C3 plasmid (No. 2)
  • Lane 4: Extraction of pSB1C3 plasmid (No. 3)
  • Lane 5: Extraction of plasmid containing lacI (No. 1)
  • Lane 6: Extraction of pSB1AK3 with BBa_B0014 plasmid containing double terminator (No. 1)
  • Lane 7: 1kb DNA ladder


(No. 1)


(No. 2)


(No. 3)


(No. 1)

Double terminator

(No. 1)

92.1 µl/ml 110.0 µl/ml 110.5 µl/ml 246.1 µl/ml 246.7 µl/ml

Table 1: Nanodrop spectrophotometer results. Table represents the amount of plasmid present in µl/ml quantity.


Bands around 3000 bp were observed in lanes 2, 3 and 4, thereby indicating the correct products. Lane 5 contain the LacI plasmid, therefore it should be bigger in size as compared to the double terminator plasmid which is in lane 6. Therefore lane 5 might not contain the LacI plasmid. From the nanodrop results, the ratio of 260:280 nm ranged from 1.7 to 1.85 and the concentration ranged from 92.1 µl/ml to 246.7 µl/ml. These results indicated that the DNA extracted have low contamination and the samples are pure.


Extraction of pSB1C3 plasmid and pSB1AK3 with BBa_B0014 plasmid containing the double terminator have been successful. As well as the addition of RNAse into the P1 buffer solved the problem of contamination.

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