Difference between revisions of "Part:BBa K2550000:Experience"

Revision as of 20:07, 16 October 2018

Applications of BBa_K2550000

Figure 1: Toehold structure implemented in the T7 Toehold LacZ biobrick that was developed in the Nupack software which confirmed the shape of the toehold sequence.

The toehold sequence, part BBa_K2550100, is 108 bp long and was obtained from the 144 first generation orthogonal toehold switches collection from the 2017 Collins paper titled Toehold Switches: De-Novo-Designed Regulators of Gene Expression. This distinct riboregulator follows the T7 promoter that is assembled in the T7 Toehold LacZ biobrick. This image above is a visual representation of this unique toehold structure.

Figure 2: The T7 Toehold LacZ biobrick transformed on a chloramphenicol and Xgal plate expressing blue pigment without trigger sequence as a result of the strong T7 promoter (Image on the left). T7 Toehold LacZ and trigger sequence transformed in a dual plasmid transformation on a carbenicillin, chloramphenicol, and Xgal plate that is expressing a blue pigment due to presence of trigger sequence (Image on the right).

T7, part BBa_I719005, is a strong constitutive promoter derived from the T7 bacteriophage that expresses proteins in the presence of T7 polymerase. It was observed that the toehold expressed a blue pigment when inoculated into Xgal and Luria Broth. Although a lighter shade than when fully induced, we hypothesize that this pigment is due to toehold leakiness as a result of the strength of the T7 promoter.

Sequencing Analysis

Figure 3: Image showing the sequencing results of the T7 Toehold LacZ construct. Annotated in a white arrow is the biobricked T7 promoter (BBa_I719005); this is followed by the biobricked toehold sequence which was used in the NuPack software to illustrate the functioning Toehold Switch (BBa_K2550100). Illustrated in the maroon arrow is the biobricked LacZ gene (BBa_I732005) sequence that justified that the one base pair wobble mutation was successful.

User Reviews

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-<i>Figure 2: The T7 Toehold LacZ biobrick transformed on a chloramphenicol and xgal plate expressing blue pigment without trigger sequence as a result of the strong T7 promoter (Image on the left). T7 Toehold LacZ and trigger sequence transformed in a dual plasmid transformation on a carbenicillin, chloramphenicol, and xgal plate that is expressing a blue pigment due to presence of trigger sequence (Image on the right).</i>
+<i>Figure 2: The T7 Toehold LacZ biobrick transformed on a chloramphenicol and Xgal plate expressing blue pigment without trigger sequence as a result of the strong T7 promoter (Image on the left). T7 Toehold LacZ and trigger sequence transformed in a dual plasmid transformation on a carbenicillin, chloramphenicol, and Xgal plate that is expressing a blue pigment due to presence of trigger sequence (Image on the right).</i>
-T7, part BBa_I719005, is a strong constitutive promoter derived from the T7 bacteriophage that expresses proteins in the presence of T7 polymerase. It was observed that the toehold expressed a blue pigment when inoculated into xgal and Luria Broth. Although a lighter shade than when fully induced, we hypothesize that this pigment is due to toehold leakiness as a result of the strength of the T7 promoter.
+T7, part BBa_I719005, is a strong constitutive promoter derived from the T7 bacteriophage that expresses proteins in the presence of T7 polymerase. It was observed that the toehold expressed a blue pigment when inoculated into Xgal and Luria Broth. Although a lighter shade than when fully induced, we hypothesize that this pigment is due to toehold leakiness as a result of the strength of the T7 promoter.
 ===Sequencing Analysis===