The BBa_K2559005 is a full-length EGFP coding part improved from BBa_l714891.
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===Usage and Biology===
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The part BBa_K2559006 has an improvement on the basic part submitted by iGEM07_Peking (BBa_l714891) which is cording the SDY_EGFP. We had added 16 bp DNA to fill the length of EGFP cording sequence in BBa_l714891 to crease the BBa_K2559005. Because, we found out a 16 bp deficiency in the EGFP starting cording region of BBa_l714891, after we checked the sequence of BBa_l714891 in NCBI.
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In order to test the function of BBa_K2559005, we design a new E.coli expression vector with our new part, BBa_K2559003, a strong E.coli endogenous promoter (PrplJ). The full length EGFP in BBa_K2559005 was driven by PrplJ, and expressed in DH10B. Attention, we also used the BBa_K2559005 in the promoter strength analysis of our other two new parts, the BBa_K2559004 and BBa_K2559011 which are weak E.coli endogenous promoters (PdapA and PcaF).
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[[File:Scau-china-2018-11.png|800px|thumb|center|Figure 1 The result of the fluorescent intensity measurement ]]
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[[File:Scau-china-2018-12.png|800px|thumb|center|Figure 2 Fluorescent intensity of BBa_K2559006 driven by by PrplJ, Pdapa and PcaiF promoter.]]
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We conclude that our improved new part, full length EGFP cording biobrick , BBa_K2559005 can work well in DH10B, and we also hoped that our improvement on the BBa_l714891 can help the future application of this part.
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To expand the application of BBa_K2559005, we search the BBa_J04450 stored in Registry. The BBa_J04450 part is a strong RFP expression vector for E.coli because of the strong RBS. As the main page BBa_J04450 mentioned, the E.coli colonies with BBa_J04450 were red in color under natural light after about 18 hour culture in LB plate. We used the BBa_K2559005 to replace the RFP region in BBa_J04450, the restructured vector from those two parts is BBa_K2559009.
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We transferred the BBa_K2559009 to DH5α by heat-shock transformation, and found that the fluorescence signal can be observe clearly under the UV.
Revision as of 06:39, 16 October 2018
SDY_EGFP
a strong reporter
measured by fluorescence microscope
[http://2018.igem.org/Team:AFCM-Egypt# Team:AFCM-Egypt 2018] provided specific characterization for this part by using eGFP for experimental characterization of lentiviral transfer plasmid transfection efficacy and apoptotic effect on colorectal cancer cell line RKO.
