Difference between revisions of "Part:BBa K1041000"

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Revision as of 11:00, 17 September 2013

RFP Coding Device

Team NRP-UEA (2013) used mutagenesis to add a Nde1 site at the start of the RFP coding region of the biobrick BBa_J04450. This enables a restriction digest with Nde1 to be performed allowing the RFP coding region to be excised from the plasmid.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 781
    Illegal AgeI site found at 893
  • 1000
    COMPATIBLE WITH RFC[1000]

Composite part of the following BioBricks:

LacI
R0010
B0034		mRFP1
E1010
B0015

Characterisation

Characterisation of this biobrick involved comparisons with the original Bba_J04450 biobrick by performing transformations, restriction digests and BLAST analysis. We also had our biobrick sequenced.

Transformation

Parts BBa_K1041000 and BBa_J04450 were transformed into Alpha-Select competent E.Coli cells and plated onto LB Agar plates which contained the antibiotic ampicillin. Both plates have colonies that appear red under natural light fig.1. Therefore the addition of a NdeI site has not effected the ability for the RFP gene to be transcribed.

Fig 1: Plates containing (left) BBa_K1041000 and (right) BBa_J04450 visualized under non-UV lightbox


Sequencing

The biobrick was sent off to a company for sequencing.

K1041000 sequencing data part 1
K1041000 sequencing data part 2
K1041000 sequencing data part 3







BLAST Analysis

The data we recieved back from the sequencing company was aligned using BLAST with the expected DNA sequence fig.2