Part:BBa_K4370010
STREPTO_E.coli_CLONING_MODULE
The part allows the easy cloning of Streptomyces genes under the control of a strong and constitutive promoter using a red/white screening of the clones of interest in E. coli used as a chassis for cloning. Therefore the module contains parts specific to Streptomyces chassis as well as parts dedicated to the expression in E. coli to perform cloning.
Notably, this part contains the Streptomyces constitutive strong promoter KasOP* followed by a RBS from of capsid protein obtained from Streptomyces temperate phage ϕC31 that has been previously described as highly efficient to promote translation when combined with the kasOP* promoter (Bai et al., PNAS, 2015). The NdeI site (catATG) allows the easy cloning of genes between NdeI (ATG/start codon) and SpeI sites (this latter site been in the suffixe). Between these two sites is inserted an cassette for the expression of mRFP1 (BBa_E1010) under the control of an E. coli promoter . This RFP expression module is followed by two stop codons and two terminators (BB0010 and B0012).
Therefore this module allows the easy red/white cloning of sequences of interest in place of the E. coli expression module. This part is designed for the cloning of Streptomyces genes of interest in place of this cassette using an easy visual screening (pink if the E. coli cassette is present in the plasmid, white if it has been replaced by the gene of interest).
This module is also designed to be easily cloned between NotI sites in the collection of Streptomyces integrative vectors published by Aubry and collaborators (AEM, 2019).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 792
Illegal AgeI site found at 904 - 1000COMPATIBLE WITH RFC[1000]
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