Part:BBa_K2043009:Design
_ bpuI-FBD10 laccase codon optimized for E.coli
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
Fabric Binding Domain 10 was fused downstream in the 5' end of the original bpul CDS
FBD 10 is a positive control obtained from literature. This 7 amino acid sequence was obtained by the phage display technique. They screened for small peptide ligands that selectively bind to the Cellulose nano whiskers.
A linker was introduced in the DNA sequence between FBD10 and the CDS of bpul
His-tag is added in the C-terminal for protein purification
The sequence was codon optimized for E. coli and restriction sites for the restriction enzymes BpiI, BsaI and BsmBI were avoided.
The sequence was confirmed by sequencing and no mutations were observed.
The construction was done by Golden Gate technique using BsaI as restriction enzyme to obtain the desired cohesive ends.
Source
Bacilus pumillus
References
"Identification and characterization of a cellulose binding heptapeptide revealed by phage display." Guo, Jing, et al., Biomacromolecules 14.6 (2013): 1795-1805.
"Screening for cellulose nanowhiskers binding peptides by phage display." Guo, Jing, et al. 2010 Pittsburgh, Pennsylvania, June 20-June 23, 2010. American Society of Agricultural and Biological Engineers, 2010.