pDawn-LKD-T1

Description

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

Usage

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

Figure 1: Genetic circuit of pDawn(cI-LVA)-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)-LKD-T1-pSEVA331 in response to blue light lysis in E. coli TOP10.

As shown in Figure 2, except for the 16^th bacterial colony,almost all of the plaque did not grow under blue light. To further show the lysis effect of pDawn(cI-LVA)-LKD-T1-TOP10, we measured the OD ₆₀₀ values of pDawn(cI-LVA)-LKD-T1-TOP10 periodically and ploting the growth curve diagram . As shown in Figure 3,pDawn(cI-LVA)-LKD-T1-TOP10 was not excellent.

Figure 2:Growth situation of pDawn(cI-LVA)-LKD-T1-TOP10 in the dark and under the blue light.

Figure 3:Growth curves diagram of pDawn(cI-LVA)-LKD-T1-TOP10 in different growth conditions.

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.