lambda lysozyme with start codon and no stop codon in BBb format. More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR al0001/al0004 on pSB3C6-Bjh1501 (505bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI, 2472+910, L) Product is pBca1256-K112302 ----------------------------------------------- al0001 Forward Biobricking of lambda lysozyme with start codon and no stop codon CCATAGAATTCatgAGATCTatggtagaaatcaataatcaacgtaaggcgttcctcg al0004 Reverse Biobricking of lambda lysozyme with start codon and no stop codon CGttaGGATCCtacatcaatctctctgaccgttccgc
T4 lysozyme gene ready to have an rbs attached in front, with a stop codon, in BBb format. ---- This part is in BBb Format. It is flanked by BamHI and BglII sites instead of XbaI and SpeI. More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR bx17/bx16 on Bjh1302 (530 bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI) Product is pBca1256-K112016 ---------------------------------------- bx17 Forward biobricking of {a~T4 Endolysin!} ccagtGAATTCatgAGATCTtattatgaatatatttgaaatgttacgtatagatg bx16 Reverse biobricking of {a~T4 Endolysin!} cgttaGGATCCtagatttttatacgcgtcccaagtgcc
T4 lysozyme gene with a start codon, without a stop codon, in BBb format. ---- This part is in BBb Format. It is flanked by BamHI and BglII sites instead of XbaI and SpeI. More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR bx13/bx16 on Bjh1302 (523 bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI) Product is pBca1256-K112014 ---------------------------------------- bx13 Forward biobricking of {T4 Endolysin>} ccagtGAATTCatgAGATCTatgaatatatttgaaatgttacg bx16 Reverse biobricking of {T4 Endolysin>} cgttaGGATCCtagatttttatacgcgtcccaagtgcc
Composite part fot TetLVA TetPromoter and T4
This part is a composite structure which contains a promoter, RBS side and and a tetracycline repressor from transposon Tn10. It is transcribed in the opposite direction for the coding sequence of the repressor AraC. Through the binding to L+arabinose, the conformation of AraC changes.This causes the protein to diffuses from the DNA thereby inducing transcription. Through the RBS, the definition of the RBS efficiency could be observed. As the second part we added Terminator+Tetr Promoter+T4 Endolysin. For this part an artifical Terminator is used (generally added to another parts). The promoter is a TetR repressible promoter.It is sequenced for pTet inverting regulator. Promoter is constitutively ON and repressed by TetR. TetR repression is inhibited by the addition of tetracycline or its analog part
L(+) - Arabinose is in sugar form and harmless The designed promoter has low background activity, so the membrane proteins will not damage the cells before induction. Bidirectional, with the reverse estimated to be more effective than the forward. Has a polyA tail of 9 residues. High expression of lysozyme only is toxic to bacteria. BBa_R0040 (TetR repressible promoter) is based on a cI promoter. It has been modified to include two TetR binding sites. Constructed by DNA synthesis.
References: //parts.igem Schleif, R. (2000). "Regulation of the L-arabinose operon of Escherichia coli." Trends Genet 16(12): 559-565. Ren, H., D. Yu, et al. (2009). "High-level production, solubilization and purification of synthetic human GPCR chemokine receptors CCR5, CCR3, CXCR4 and CX3CR1 http://openwetware.org/wiki/Titratable_control_of_pBAD_and_lac_promoters_in_individual_E._coli_cells#pBAD_promotersOpenWetWare http://www.ncbi.nlm.nih.gov/pubmed/7768852?dopt=Abstract Enterobacteria phage T4 http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=29366675&from=66503&to=66997&view=gbwithparts modified E. coli thr terminator, replaced all A-T pairs in stem with C-G pairs BiblioPlus Extension Error fetching PMID 9092630:
lambda lysozyme ready to have an rbs attached with a stop codon More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR al0005/al0002 on pSB3C6-Bjh1501 (512bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI, 2472+910, L) Product is pBca1256-K112304 ----------------------------------------------- al0005 Forward Biobricking of lambda lysozyme with spacer CCATAGAATTCatgAGATCTgaagatggtagaaatcaataatcaacgtaaggcgttcctcg al0002 Reverse Biobricking of lambda lysozyme with spacer CGttaGGATCCtcatacatcaatctctctgaccgttccgcc
Lambda Lysozyme with stop codon and no start codon in BBb format
PCR al0003/al0002 on pSB3C6-Bjh1501 (505bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI, 2472+910, L) Product is pBca1256-K112301 ----------------------------------------------- al0003 Forward Biobricking of Lambda Lysozyme with stop codon and no start codon CCATAGAATTCatgAGATCTgtagaaatcaataatcaacgtaaggcgttcctcg al0002 Reverse Biobricking of Lambda Lysozyme with stop codon and no start codon CGttaGGATCCtcatacatcaatctctctgaccgttccgcc
Truncated lysostaphin coding sequence. Lysostaphin has has a specific lytic action against S.aureus.
Lysostaphin is a zinc metalloenzyme that has a specific lytic action against S.aureus. Lysostaphin has activities of three enzymes namely, endo-β-N-acetyl glucosamidase, N-acteyl-muramyl-L-alanine amidase, glycylglycine endopeptidase. Glycylglycine endopeptidase lyses staphylococcal cells by hydrolyzing glycylglycine bonds in the poly-glycine bridges which form cross links between glycopeptide chains in cell wall peptidoglycan of S.aureus cells. The wild-type lysostaphin gene encodes a preproenzyme, and the conversion of prolysostaphin to mature lysostaphin occurs extracellularly and involves the removal of the hydrophilic tandem repeat portion of the proenzyme. In order to directly produce mature lysostaphin, we truncate the preprolysostaphin and prolysostaphin sequence.
In order to directly produce mature lysostaphin, we truncate the preprolysostaphin and prolysostaphin sequence.
T4 lysozyme gene with start and stop codons, in BBb format. ---- This part is in BBb Format. It is flanked by BamHI and BglII sites instead of XbaI and SpeI. More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR bx13/bx14 on Bjh1302 (526 bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI) Product is pBca1256-K112012 ---------------------------------------- bx13 Forward biobricking of {T4 Endolysin!} ccagtGAATTCatgAGATCTatgaatatatttgaaatgttacg bx14 Reverse biobricking of {T4 Endolysin!} cgttaGGATCCttatagatttttatacgcgtcccaagtgcc
T4 lysozyme gene without a start codon, with a stop codon, in BBb format. ---- This part is in BBb Format. It is flanked by BamHI and BglII sites instead of XbaI and SpeI. More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR bx15/bx14 on Bjh1302 (523 bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI) Product is pBca1256-K112013 ---------------------------------------- bx15 Forward biobricking of {<T4 Endolysin!} ccagtGAATTCatgAGATCTaatatatttgaaatgttacgtatagatg bx14 Reverse biobricking of {<T4 Endolysin!} cgttaGGATCCttatagatttttatacgcgtcccaagtgcc
The lysozyme from enterobacteria phage T4 degrades peptidoglycan layer.
High expression of lysozyme only is toxic to bacteria.
Enterobacteria phage T4 http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=29366675&from=66503&to=66997&view=gbwithparts
T4 endolysin under the control of T7 promotor
Ane Fernanda Beraldi Zeidler, Luige A. Llerena Calder??n and Marcos Henrique de Moraes
This construction is a device composed of: BBa_I746911 - BBa_K112806 - Bba_B0014. This device contain the T4 endolysin, a lysozyme from enterobacteria phage, that degrades peptidoglycan layer, under the control of the T7 promoter.
For more accurate tests about the endolysin effectiveness, this part has to be under the control of an inducible promoter, like the T7 promoter.
This device is a junction of previously designed parts: BBa_I746911 - Designed by Stefan Milde Group: iGEM07_Cambridge (2008-09-30) BBa_K112806 - Designed by Jin Huh Group: iGEM08_UC_Berkeley (2008-10-31) Bba_B0014 - Designed by Reshma Shetty Group: Registry (2003-07-16)
Tlanslational unit of Colicin Lysis Protein (for colicin-E3)
Colicin lysis protein allows colicins to be released. The mechanism how the colicin lysis protein allows colicin release has not been fully elucidated, but it is sure that this protein raise membrane permiability and cause quasilysis.[1] Colicins are a cytotoxins which are released to environment and kill other related strains.
This part causes cell death. It contains holin/antiholin, endolysin and Rz/Rz1 genes.
We made point mutation and prepared wild type phage DNA.
λ -Hind Ⅲ digest (product of TaKaRa Bio)
A DNA cassette that undergo homologous recombination into E.coli genome that contains homologous regions at each end, Kanamicin resistance gene (KanR) and death operon containing Lambda holin and lysozime genes and controlled by a Lambda cI regulated promoter
part is synthetic. holin and lysozime originates from lambda bacteriophage. the homology regions were taken from E.coli BL21 genome
atgagtaaag gagaagaact tttcactgga gtggtcccag ttcttgttga attagatggc gatgttaatg ggcaaaaatt ctctgtcagt ggagagggtg aaggtgatgc aacatacgga aaacttaccc ttaattttat ttgcactact gggaagctac ctgttccatg gccaacactt gtcactactt tctcttatgg tgttcaatgc ttctcaagat acccagatca tatgaaacag catgactttt tcaagagtgc catgcccgaa ggttatgtac aggaaagaac tatattttac aaagatgacg ggaactacaa gacacgtgct gaagtcaagt ttgaaggtga tacccttgtt aatagaatcg agttaaaagg tattgatttt aaagaagatg gaaacattct tggacacaaa atggaataca actataactc acataatgta tacatcatgg gagacaaacc aaagaatggc atcaaagtta acttcaaaat tagacacaac attaaagatg gaagcgttca attagcagac cattatcaac aaaatactcc aattggcgat ggccctgtcc ttttaccaga caaccattac ctgtccacac aatctgccct ttccaaagat cccaacgaaa agagagatca catgatcctt cttgagtttg taacagctgc taggattaca catggcatgg atgaactata caaa
Mutated endolysin specific for Acinetobacter baumannii.
Roger Milton Rubio S??nchez, Enrique Chimal, Enrique Palomino, Brian Morteo
This biobrick includes the coding sequence for a mutated version (PlyAB1) of lysAB2 which specifically targets Acinetobacter baumannii. This protein was mutated in 5 aminoacids to promote its specific binding to the baumannii species.
This sequence was optimised for its usage in RFC10 standard and in E. coli as chassis.
lambda lysozyme with start and stop codons in Bbb format
T4 lysozyme without start or stop codons, in BBb format. ---- This part is in BBb Format. It is flanked by BamHI and BglII sites instead of XbaI and SpeI. More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR bx15/bx16 on Bjh1302 (520 bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI) Product is pBca1256-K112015 ---------------------------------------- bx15 Forward biobricking of {<T4 Endolysin>} ccagtGAATTCatgAGATCTaatatatttgaaatgttacgtatagatg bx16 Reverse biobricking of {<T4 Endolysin>} cgttaGGATCCtagatttttatacgcgtcccaagtgcc
lambda lysozyme with no start/stop codon in BBb format More information about the BBb Format is available at: [http://openwetware.org/wiki/Template:AndersonLab:BBb_Standard BBb Standard Description Page]
PCR al0003/al0004 on pSB3C6-Bjh1501 (502bp, EcoRI/BamHI/DpnI) Sub into pBca1256 (EcoRI/BamHI, 2472+910, L) Product is pBca1256-K112303 ----------------------------------------------- al0003 Forward Biobricking of lambda lysozyme with no start/stop codon CCATAGAATTCatgAGATCTgtagaaatcaataatcaacgtaaggcgttcctcg al0004 Reverse Biobricking of lambda lysozyme with no start/stop codon CGttaGGATCCtacatcaatctctctgaccgttccgc
Endolysin is a lysozyme which cleaves the glycosidic bond between the C-1 of N-acetylmuramic acid (NAM) and the C-4 of N-acetylglucosamine (NAG) in the bacterial peptidoglycan (PG). This leads to the complete lysis of the cell. Holin is a protein that assembles together to form pores on inner membrane of bacteria allowing lysozyme to reach periplasm and degrade peptidoglycan layer. It is sourced from T4 bacteriophage. This composite part can be used to induce self-destruction with a regulatory device in the upstream region.
There is no RBS in the upstream region, so during the assemble procedure, this must be taken care of.
Gene sequences of endolysin and holin are gathered from the Registry.
Putative lysin from the Bordetella phage genome. Contains sequence similarity to ElmA, a protein in E. coli that breaks down capsid polysaccharides.
Isolated from phage genome, made BioBrick-compatible.
PCR-amplification from isolated BPP phage genome. Described as a putative lysin in: M Liu, M Gingery, SR Doulatov, Y Liu, A Hodes, S Baker, P Davis, M Simmonds, C Churcher, K Mungall, MA Quail, A Preston, ET Harvill, DJ Maskell, FA Eiserling, J Parkhill, JF Miller. (2004). Genomic and genetic analysis of Bordetella bacteriophages encoding reverse transcriptase-mediated tropism-switching cassettes. Journal of Bacteriology 186(5):1503.
This part comes from a Bacteriophage Holin Gene pS105 This is a variation of the S gene of the "Cell Lysis Cassette." When it is paired with an activator, the cassette is expressed and cell lysis is induced. The S105 gene has the initial 6 base pairs at the beginning of the wild type S gene sequence removed. This causes cell lysis to be induced at a faster rate than the wild type. The added Terminator is artificial and large.
No design considerations during Holin Gene Bidirectional, with the reverse estimated to be more effective than the forward. Has a polyA tail of 9 residues.
Brown iGEM Team by Ry Young at Texas A&M Modified E. coli thr terminator, replaced all A-T pairs in stem with C-G pairs