pDawn(cl)-X174E-T1

Description

This composite part is a generator consisting of pDawn(cI) (BBa_K4325010), X174E (BBa_K1835500) and T1 terminator (BBa_K3033016).

Usage

We inserted the blue light responsive system pDawn(cI) (BBa_K4325010) and the lysis gene X174E (BBa_K1835500) into the pSEVA331 expression vector, which was introduced into E. coli TOP10. The bacterial colonies that grew in the dark but did not grow under blue light were screened to verify the responsiveness of pDawn(cI) to blue light.

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(cl)-X174E-T1 by evaluating the response to blue light.

Figure1. (1) pDawn(cI)-X174E-T1; (2) pDawn(cI-LVA)-X174E-LVA-T1; (3) pDawn(cI)-LKD-T1; (4) pDawn(cI-LVA)-RBS070-LKD-LVA-T1 were constructed successfully.

Figure2. Growth condition of E.coli TOP10-pDawn(cI)-X174E-T1 in the dark and under blue light.

As shown in Figure 1, we successfully constructed pDawn(cI)-X174E-T1 and transformed it into E. coli TOP10, selecting bacteria colonies and culturing in different growth conditions. As shown in Figure 2, the 9^th colony of E.coli TOP10-pDawn(cI)-X174E-T1 grew in the dark, but did not grow under blue light.

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.

[2] Witte, A., Wanner, G., Sulzner, M. & Lubitz, W. Dynamics of PhiX174 protein E-mediated lysis of Escherichia coli. Arch. Microbiol. 157, 381-8 (1992).