Difference between revisions of "Part:BBa K4325026"
Line 6: Line 6:
 
===Usage===
 
===Usage===
 
<p>The pDawn(cI-LVA) blue light response system (<partinfo>BBa_K1075044</partinfo>) and lysis gene LKD (<partinfo>BBa_K4325004</partinfo>) were inserted into the pSEVA331 expression vector, which was inserted into <i>E. coli</i> TOP10, screened out the bacterial colony which grew in the dark but did not grow under blue light. Finally, the pDawn(cI-LVA)-RBS070-LKD-T1 plasmid was selected and inserted into <i> G. hansenii</i> ATCC53582 by electroporation to verify the responsiveness of pDawn(cI-LVA) under blue light.</p>
 
<p>The pDawn(cI-LVA) blue light response system (<partinfo>BBa_K1075044</partinfo>) and lysis gene LKD (<partinfo>BBa_K4325004</partinfo>) were inserted into the pSEVA331 expression vector, which was inserted into <i>E. coli</i> TOP10, screened out the bacterial colony which grew in the dark but did not grow under blue light. Finally, the pDawn(cI-LVA)-RBS070-LKD-T1 plasmid was selected and inserted into <i> G. hansenii</i> ATCC53582 by electroporation to verify the responsiveness of pDawn(cI-LVA) under blue light.</p>
−
[[File:K26 11.png|600px|thumb|center|Figure 1: <p></p>Gene circuit of pDawn(cI-LVA)-RBS070-LKD-T1.]]
+
[[File:K26 11.png|600px|thumb|center|Figure 1: Gene circuit of pDawn(cI-LVA)-RBS070-LKD-T1.]]
  
 
===Sequence and Features===
 
===Sequence and Features===

Revision as of 17:47, 11 October 2022

pDawn-RBS070-LKD-T1

Description

This composite part is a generator containing pDawn(cI-LVA) (BBa_K1075044) and LKD (BBa_K4325004).

Usage

The pDawn(cI-LVA) blue light response system (BBa_K1075044) and lysis gene LKD (BBa_K4325004) were inserted into the pSEVA331 expression vector, which was inserted into E. coli TOP10, screened out the bacterial colony which grew in the dark but did not grow under blue light. Finally, the pDawn(cI-LVA)-RBS070-LKD-T1 plasmid was selected and inserted into G. hansenii ATCC53582 by electroporation to verify the responsiveness of pDawn(cI-LVA) under blue light.

Figure 1: Gene circuit of pDawn(cI-LVA)-RBS070-LKD-T1.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 2171
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 63
    Illegal NgoMIV site found at 195
    Illegal NgoMIV site found at 289
    Illegal NgoMIV site found at 582
    Illegal NgoMIV site found at 1076
    Illegal NgoMIV site found at 1094
    Illegal NgoMIV site found at 1184
    Illegal AgeI site found at 414
    Illegal AgeI site found at 1542
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1643
    Illegal BsaI.rc site found at 525


2022 SZPT-China

Characterization

1.Batch screening of pDawn(cI-LVA)-RBS070-LKD-T1 in response to blue light lysis in E. coli TOP10.

As shown in Figure 2,except for the 11th and 13th bacterial colony,almost all of the bacterial colony did not grow under blue light. To further explore the lysis effect of pDawn(cI-LVA)-RBS070-LKD-T1-TOP10, we measured the OD 600 values of pDawn(cI-LVA)-RBS070-LKD-T1-TOP10 and ploting the growth curve diagram. As shown in Figure 3, the pDawn (cI-LVA)-RBS070-LKD-T1-TOP10 lysis effect was well in the first eight hours, but was unstable after fifteen hours.

Figure 2: Growth conditions of pDawn(cI-LVA)-RBS070-LKD-T1-TOP10 in the dark and under the light.
Figure 3: Growth curves diagram of pDawn(cI-LVA)-RBS070-LKD-T1-TOP10.


References

[1]Robert Ohlendorf, Roee R. Vidavski, Avigdor Eldar et.al.From Dusk till Dawn: One-Plasmid Systems for Light-Regulated Gene Expression.Journal of Molecular Biology, 08 Jan 2012, 416(4):534-542.