Difference between revisions of "Part:BBa K3714004"
Line 12: Line 12:
 
<partinfo>BBa_K3714004 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K3714004 SequenceAndFeatures</partinfo>
 
<hr/>
 
<hr/>
−
By introducing mutations in loop Ⅰ and loop Ⅱ[1], we achieved an aptazyme with higher responsing fold and less leakage under high theophylline concentration (Fig.1).  
+
By introducing mutations in loop Ⅰ and loop Ⅱ[1], we achieved an aptazyme with higher responsing fold and less leakage under high theophylline concentration (Fig.1). We characterized the responding fold of these aptazymes under diffrent theophylline concentrations.  
 
<center>[[File:K3714004-1.jpg]]</center>
 
<center>[[File:K3714004-1.jpg]]</center>
  

Revision as of 23:50, 21 October 2021


Improved theophylline biosensor

An improved aptazyme responsing to theophylline, which means its self-cleaving activity can be inhibited by adding theophylline. This aptazyme shows higher sensibility than an existing part (BBa_J119449).

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
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

By introducing mutations in loop Ⅰ and loop Ⅱ[1], we achieved an aptazyme with higher responsing fold and less leakage under high theophylline concentration (Fig.1). We characterized the responding fold of these aptazymes under diffrent theophylline concentrations.

K3714004-1.jpg
Figure 1 | Improvement of an existing theophylline aptazyme. Cleavage properties of two aptazymes under different theophylline concentration. The aptazymes were transcribed in vitro with certain amount of theophylline, and the products were run a a denatured PAGE. The number above each lane means dilution fold of saturated theophylline aqueous solution at 25℃.

To further quantify the outcome of our engineering, we used a lower concentration gradient. Our aptazyme exhibits a 2-fold change of cleavage proportion at ~1mg/ml (0.125x dilution of saturated solution at room temperature) while the origin apatazyme (BBa_J119449) shows only 1.3-fold change (Fig.2).

K3714004-2.jpg
Figure 2 | Improvement of an existing theophylline aptazyme. a.Cleavage properties of two aptazymes under a lower theophylline concentration gradient. b. A diagram showing the intensity percentage of the cleaved band of each lane in a. Intensity was measured by ImageJ.