Promoter chiA74 from Bacillus thuringiensis (Ba_K2057000)
The promoter chiA74 (chiA74p) controls the expression of the endochitinase
chiA74 from a strain of Bacillus thuringiensis. This regulatory element is also useful
to drive expression of a gene in Escherichia coli in a constitutive form (Barboza-
Corona et al. 2003, 2014). We have shown that chiA74p drive the expression of
the green fluorescent protein (gfp) gene in E. coli. In this sense, chiA74p can be
used to allow the synthesis of a gene in E. coli that want to be expressed
constitutively. Biobrick also include the ribosome binding sequence of chiA74. This
is one of the promoter that our team will use to express constitutively the lasR gene
in E. coli. Promoter chiA74p has the consensus sequences in 374-384 bp (-35)
region) and 399-404 (-10). The ribosome-binding site is located on 559-564 bp. (II)
The whole sequence of the chiA74 gene including its regulatory elements
(promoter, RBS, transcriptional terminator) is found under accession number
AF424979 (GenBank nucleotide sequence database).
References
Barboza-Corona JE, Delgadillo-Ángeles JL, Castañeda-Ramíırez JC, Barboza-
Pérez UE, Casados-Vázquez LE, Bideshi DK and del Rincón-Castro MC (2014)
Bacillus thuringiensis subsp. kurstaki HD1 as a factory to synthesize alkali-labile
ChiA74∆sp chitinase inclusions, Cry crystalsand spores for applied use. Microb
Cell Fact 13, 15. doi: 10.1186/1475-2859-13-15
Barboza Corona JE, Nieto-Mazzocco E, Velázquez-Robledo R, Salcedo-
Hernandez R, Bautista M, Jiménez B, Ibarra JE (2003) Cloning, sequencing and
expression of the chitinase chiA74 from Bacillus thuringiensis. Appl Environ
Microbiol. 2003 Feb;69(2):1023-9.
Promoter BtI-BtII (Ba_K2057001)
The BtI-BtII promoter (BtI-BtIIp) is a dual strong promoter that controls the
expression of Cry1Ac proteins in Bacillus thuringiensis, but it allows the expression
of a gene constitutively in Escherichia coli. In this sense, BtI-BtIIp can be used to
allow the synthesis of a gene in E. coli that want to be expressed constitutively in
E. coli. This is one of the promoter that our team will use to express constitutively
the lasR gene in E. coli. The biobrick also include a ribosome-binding site to to
start the synthesis of a proteins. This promoter can can be used not only in E. coli
but also in B. subtilis and B. thuringiensis. Promoter BtI-BtII has the consensus
sequences in 285-291 bp (BtI, -35 region), 294-302 (Bt II, -10) and 306-313 (BtI, -
10). The ribosome binding site is located on 377-381 bp. The whole sequence of
the cry1Ac gene including its regulatory elements (promoter, RBS, transcriptional
terminator) is found under accession number M35524 (GenBank nucleotide
sequence database).
References
Von Tersch MA, Robbins HL, Jany CS, Johnson TB (1991) Insecticidal toxins
from Bacillus thuringiensis subsp. kenyae: gene cloning and characterization and
comparison with B. thuringiensis subsp. kurstaki CryIA(c) toxins Appl. Environ.
Microbiol. 57 (2): 349-358.
Sedlak M1, Walter T, Aronson A (2000) Regulation by overlapping promoters of
the rate of synthesis and deposition into crystalline inclusions of Bacillus
thuringiensis delta-endotoxins. J Bacteriol. 182(3):734-41.
Alginate lyase from Bacillus thuringiensis (BBa_K2057002)
Alginate lyases destroy the alginate, which is the main component of biofilm in
many bacteria, including that produced by Pseudomonas aeruginosa. As it has
never reported the cloning of alginate lyase gene in B. thuringiensis we designed
the primers using the genome of Bacillus thuringiensis israelensis AM65-52
(GenBank accession number: CP013275.1). We amplified the alginate lyase gen
from the genome of Bacillus thuringiensis 4Q7. We are in process of sequencing
and test its functionality in E. coli
References
Farrel EK, Tipton PA (2012) Functional characterization of AlgL, an alginate
lyase from Pseudomona aeruginosa. Biochem. 51:10259-10266.
Tielen, P, Kuhn H, Rosenau F, Jaeger KE, Flemming HC, Wingender J (2013)
Interaction between extracellular lipase LipA and the polysaccharide alginate of
Pseudomonas aeruginosa. BMC Microbiol. 159: 221–228.
Signal peptide of ChiA74 (BBa_K2057007)
Signal peptide of ChiA74 translocate chitinase ChiA74 in Bacillus thuringiensis and
Escherichia coli. Sequence comes from the endochitinase gene chiA74 that is
located on a chromosomal sequence of B. thuringiensis. It will be used to
translocate the alginate lyase and the microcin in E. coli.
References
Barboza-Corona JE, Delgadillo-Ángeles JL, Castañeda-Ramíırez JC, Barboza-
Pérez UE, Casados-Vázquez LE, Bideshi DK and del Rincón-Castro MC (2014)
Bacillus thuringiensis subsp. kurstaki HD1 as a factory to synthesize alkali-labile
ChiA74∆sp chitinase inclusions, Cry crystalsand spores for applied use. Microb
Cell Fact 13, 15. doi: 10.1186/1475-2859-13-15
Barboza Corona JE, Nieto-Mazzocco E, Velázquez-Robledo R, Salcedo-
Hernandez R, Bautista M, Jiménez B, Ibarra JE (2003) Cloning, sequencing and
expression of the chitinase chiA74 from Bacillus thuringiensis. Appl Environ
Microbiol. 2003 Feb;69(2):1023-1029.
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