Difference between revisions of "Part:BBa K3431051"

Revision as of 09:15, 25 October 2020 (view source) IrisRChen (Talk | contribs) ← Older edit		Revision as of 09:15, 25 October 2020 (view source) IrisRChen (Talk | contribs) (→‎Characterization) Newer edit →
Line 20:		Line 20:
	</html>		</html>
−	<br><br>	+	<br>
	<b>Results</b> As shown above, glucose concentration increased with time in the ON state but remained low in the OFF state. The result suggested the regulatory function of the toehold switch.		<b>Results</b> As shown above, glucose concentration increased with time in the ON state but remained low in the OFF state. The result suggested the regulatory function of the toehold switch.

---

Revision as of 09:15, 25 October 2020

Banana_ToeholdSwitch-Regulated Invertase

Introduction

Banana_ToeholdSwitch-Regulated Invertase is a genetic device consisting of 4 basic parts: T7 promoter (BBa_I719005), banana toehold switch (BBa_K3431039), Thermotoga maritima Invertase (BBa_K3431000), and T7 terminator (BBa_K731721). The protein expression of this part is regulated by the RNA of the banana.

Components

As for the T7 promoter (BBa_I719005) and T7 terminator (BBa_K731721), they are the essential genetic elements for the PURExpress protein synthesis kit. The mechanism of the detection is mainly based on the regulatory part, Banana Toehold Switch (BBa_K3431039). Its sequence is from a modular synthetic biology education kit, banana toehold sensor sfGFP (Biobits) 1 . Upon binding with banana RNA, its hairpin structure can be opened up and the ribosomes can bind with its RBS (ribosome binding site), triggering the translation process of the downstream reporter, invertase (BBa_K3431000). The reporter invertase can convert sucrose into glucose, which can be easily measured by a personal glucose meter (PGM) Therefore, the part can be used as a detection tool.

Characterization

To further understand its functionality, 2020 iGEM CSMU-Taiwan conducted a test on it. The plasmid would be transcribed and translated with the protein synthesis kit at 37℃ for 2 hours. We would then add 5μl of 0.5M sucrose and measured the glucose concentration with the Rightest TM GS550 glucose meter every 10 minutes at 55℃ for an hour. In our experiment, the ON state refers to the conditions with miRNA triggers; while the OFF state means that there was no miRNA in the environment.

Figure 1. The glucose concentration in 60 minutes invertase enzymatic reaction. The green line refers to the conditions with banana triggers, while the blue line refers to the conditions without a banana trigger.

Results As shown above, glucose concentration increased with time in the ON state but remained low in the OFF state. The result suggested the regulatory function of the toehold switch.

Sequence and Features