Difference between revisions of "Part:BBa K1694014"
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<h1>'''Introduction:'''</h1> | <h1>'''Introduction:'''</h1> | ||
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To display the antibody outside the E.coli, we used Lipoprotein-Outer membrane protein A (Lpp-OmpA). According to the paper reference [1], We chose the first 9 amino acid of Lpp, and the 46~159 amino acid of OmpA. | To display the antibody outside the E.coli, we used Lipoprotein-Outer membrane protein A (Lpp-OmpA). According to the paper reference [1], We chose the first 9 amino acid of Lpp, and the 46~159 amino acid of OmpA. | ||
<br> | <br> | ||
In order to change the scFv parts easily, we added a NcoI restriction site between OmpA and scFv so that we can change vario us scFv DNA sequence using the NcoI restriction enzyme. | In order to change the scFv parts easily, we added a NcoI restriction site between OmpA and scFv so that we can change vario us scFv DNA sequence using the NcoI restriction enzyme. | ||
+ | [[File:ompascfv.png|400px|thumb|center|'''Fig.2''' ompascfv]] | ||
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<h1>'''Experiment:'''</h1> | <h1>'''Experiment:'''</h1> | ||
+ | [[File:OC.png|200px|thumb|left|'''Fig.3'''The PCR result of the Lpp-OmpA-N+scFv. The DNA sequence length is around 1000~1200 bp, so the PCR products should appear at 1200~1400 bp.]] | ||
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After receiving the DNA sequences from the gene synthesis company, we recombined each OmpA-N-scFv gene to PSB1C3 backbones and conducted a PCR experiment to check the size of each of the OmpA-N-scFvs. The DNA sequence length of the OmpA-N-scFvs are around 1000~1200 bp. In this PCR experiment, the OmpA-N-scFv products size should be near at 1200~1400 bp. The Fig. showed the correct size of the scFv, and proved that we successful ligated the scFv sequence onto an ideal backbone. | After receiving the DNA sequences from the gene synthesis company, we recombined each OmpA-N-scFv gene to PSB1C3 backbones and conducted a PCR experiment to check the size of each of the OmpA-N-scFvs. The DNA sequence length of the OmpA-N-scFvs are around 1000~1200 bp. In this PCR experiment, the OmpA-N-scFv products size should be near at 1200~1400 bp. The Fig. showed the correct size of the scFv, and proved that we successful ligated the scFv sequence onto an ideal backbone. | ||
Revision as of 15:01, 19 September 2015
OmpA-scFv(Anti-EGFR)
Introduction:
To display the antibody outside the E.coli, we used Lipoprotein-Outer membrane protein A (Lpp-OmpA). According to the paper reference [1], We chose the first 9 amino acid of Lpp, and the 46~159 amino acid of OmpA.
In order to change the scFv parts easily, we added a NcoI restriction site between OmpA and scFv so that we can change vario us scFv DNA sequence using the NcoI restriction enzyme.
Experiment:
After receiving the DNA sequences from the gene synthesis company, we recombined each OmpA-N-scFv gene to PSB1C3 backbones and conducted a PCR experiment to check the size of each of the OmpA-N-scFvs. The DNA sequence length of the OmpA-N-scFvs are around 1000~1200 bp. In this PCR experiment, the OmpA-N-scFv products size should be near at 1200~1400 bp. The Fig. showed the correct size of the scFv, and proved that we successful ligated the scFv sequence onto an ideal backbone.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 388
- 1000COMPATIBLE WITH RFC[1000]