Difference between revisions of "Part:BBa K3866032"
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In order to construct this part, we replaced the native signal peptide with the N20 signal peptide sequence which is: 5’-atggaaggcaacacccgcgaagataactttaaacatctgctgggcaacgataacgtgaaacgcccgagcgaagcg-3’. The coding sequence was domesticated. We removed BsmBI, BsaI, BtgZI, BpiI sites in order to be compatible with GoldenBraid and MoClo. The sequence is present in pUPD2 <bbpart>BBa_K3505007</bbpart> and has overhangs compatible for GoldenBraid cloning. The CDS has position B3-B5. | In order to construct this part, we replaced the native signal peptide with the N20 signal peptide sequence which is: 5’-atggaaggcaacacccgcgaagataactttaaacatctgctgggcaacgataacgtgaaacgcccgagcgaagcg-3’. The coding sequence was domesticated. We removed BsmBI, BsaI, BtgZI, BpiI sites in order to be compatible with GoldenBraid and MoClo. The sequence is present in pUPD2 <bbpart>BBa_K3505007</bbpart> and has overhangs compatible for GoldenBraid cloning. The CDS has position B3-B5. | ||
| + | [[Image: T--Thessaly--GB-AATG-GCTT.jpeg|800px|thumb|none|<i><b>Figure 1.</b>The overhangs of this part in the GoldenBraid Grammar</i>]] | ||
| + | |||
===Verification of cloning=== | ===Verification of cloning=== | ||
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===References=== | ===References=== | ||
| − | Gao, D., Wang, S., Li, H., Yu, H., & Qi, Q. (2015). Identification of a heterologous cellulase and its N-terminus that can guide recombinant proteins out of Escherichia coli. Microbial cell factories, 14, 49. | + | Gao, D., Wang, S., Li, H., Yu, H., & Qi, Q. (2015). Identification of a heterologous cellulase and its N-terminus that can guide recombinant proteins out of Escherichia coli. Microbial cell factories, 14, 49. |
| − | Yang, M., Luoh, S., Goddard, A., Reilly, D., Henzel, W., & Bass, S. (1996). The bgIX gene located at 47.8 min on the Escherichia coll chromosome encodes a periplasmic -glucosidase. Microbiology, 142(7), 1659-1665. | + | Yang, M., Luoh, S., Goddard, A., Reilly, D., Henzel, W., & Bass, S. (1996). The bgIX gene located at 47.8 min on the Escherichia coll chromosome encodes a periplasmic -glucosidase. Microbiology, 142(7), 1659-1665. |
Revision as of 14:40, 27 September 2021
N20bglΧ GB compatible with B3-B5
This is the E. coli β-glucosidase gene bglX with N20 signal peptide instead of its native signal peptide.
Usage and Biology
Bglx belongs to the family of β-D-Glucosidases. These enzymes catalyze the hydrolysis of cellobiose (1,4-β-D-glucopyranosyl-D-glucopyranose) into two molecules of glucose. The bgIX gene is located adjacent to the dld gene at 47.8 min or 2225 kb on the E. coli chromosome. The presence of a 20 amino acids signal peptide (at the N-terminal) is consistent with the periplasmic location of the mature protein. The replacement of native signal peptide with N20- signal peptide is expected to make the mature protein an extracellular product.
Design Notes
In order to construct this part, we replaced the native signal peptide with the N20 signal peptide sequence which is: 5’-atggaaggcaacacccgcgaagataactttaaacatctgctgggcaacgataacgtgaaacgcccgagcgaagcg-3’. The coding sequence was domesticated. We removed BsmBI, BsaI, BtgZI, BpiI sites in order to be compatible with GoldenBraid and MoClo. The sequence is present in pUPD2 BBa_K3505007 and has overhangs compatible for GoldenBraid cloning. The CDS has position B3-B5.
Verification of cloning
Source
Synthesized by IDT.
Experimental Use and Experience
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 1637
Illegal AgeI site found at 1859
Illegal AgeI site found at 2048 - 1000COMPATIBLE WITH RFC[1000]
References
Gao, D., Wang, S., Li, H., Yu, H., & Qi, Q. (2015). Identification of a heterologous cellulase and its N-terminus that can guide recombinant proteins out of Escherichia coli. Microbial cell factories, 14, 49. Yang, M., Luoh, S., Goddard, A., Reilly, D., Henzel, W., & Bass, S. (1996). The bgIX gene located at 47.8 min on the Escherichia coll chromosome encodes a periplasmic -glucosidase. Microbiology, 142(7), 1659-1665.