LOCUS       sequence-204360-        6011 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  plasmid for expression of computationally designed colicin 
            endonuclease and immunity pair, des18.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6011)
  AUTHORS   Netzer R, Listov D, Lipsh R, Dym O, Albeck S, Knop O, Kleanthous C, 
            Fleishman SJ
  TITLE     Ultrahigh specificity in a network of computationally designed 
            protein-interaction pairs.
  JOURNAL   Nat Commun. 2018 Dec 11;9(1):5286. doi: 10.1038/s41467-018-07722-9.
  PUBMED    30538236
REFERENCE   2  (bases 1 to 6011)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-12-11"; volume: "9(1):5286. doi"; pages: " 
            10.1038/s41467-018-07722-9"
COMMENT     Sequence Label: pET21d-colEdes18/Imdes18
FEATURES             Location/Qualifiers
     source          1..6011
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      19..607
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     508..527
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     761..778
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    793..932
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(858..877)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1018..1040
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1034..1225)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     3077..3096
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3116..3193)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3399..3418
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3502..3521
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3502..3520
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3521..3545
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             3560..3582
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4245..4262
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4315..4333)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4329..4376
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4413..4868
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4500..4519)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4710..4731
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4894..4998
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4999..5067,5068..5859)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5217..5236)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
       61 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
      121 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
      181 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
      241 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
      301 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
      361 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
      421 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
      481 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
      541 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
      601 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
      661 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
      721 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
      781 agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
      841 caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc
      901 cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg
      961 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     1021 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt
     1081 aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca
     1141 gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa
     1201 gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg
     1261 gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac
     1321 atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc
     1381 agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac
     1441 agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact
     1501 tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg
     1561 tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct
     1621 gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca
     1681 tgcgcacccg tggggccgcc atgccggcga taatggcctg cttctcgccg aaacgtttgg
     1741 tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg
     1801 acaggccgat catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg
     1861 ctgccggcac ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga
     1921 tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg
     1981 gtcgagatcc cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc
     2041 ccgctttcca gtcgggaaac tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     2101 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     2161 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     2221 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     2281 tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact
     2341 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     2401 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     2461 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     2521 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     2581 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     2641 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     2701 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     2761 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     2821 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     2881 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     2941 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     3001 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     3061 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     3121 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc
     3181 attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag
     3241 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag
     3301 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     3361 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg
     3421 ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc
     3481 gagatctcga tcccgcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca
     3541 attcccctct agaaataatt ttgtttaact ttaagaagga gatataccat ggagagtaaa
     3601 cggaataagc cagggaaggc gacaggtaaa ggtaaaccag ttggtgaccg ttggctggac
     3661 gacgcgggta aagattctgg tgcgccgatc ccggatcgta ttgcggacaa actgcgtgac
     3721 aaagaattca aaaacttcga cgacttccgt aagaaattct gggaagaagt ttctaaagac
     3781 ccagacctgg cgaaacagtt caaacgtgcg aaccgtaacc gtattaagca gggtgaagcg
     3841 ccgttcgctc cgcaaaaaga ccaggttggc ggtcgcaaaa ctttcgaact gcaccacgac
     3901 aaaccgatct ctcaggacgg tggtgtttac gatatgaaca acatcagagt gaccacacct
     3961 aagcgacata ttgatattca tcggggtaag taaaaatgga actgaaacat agtattagtg
     4021 attataccga ggctgaattt ctggagtttg ttaagaaaat cgttgaaggt ctgctgggtc
     4081 agccgtctgt tgaagaattc atccgtctga ccgaacaccc gtctggttct gacctgatct
     4141 actacccgcg tgacgaccgt gaagactctc cggaaggtat cgttaaagaa atcaaagaat
     4201 ggcgagctgc taacggtaag tcaggattta aacagggcct cgagcaccac caccaccacc
     4261 actgagatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg
     4321 agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga
     4381 aaggaggaac tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
     4441 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     4501 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4561 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4621 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     4681 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     4741 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     4801 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     4861 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     4921 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     4981 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     5041 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     5101 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     5161 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     5221 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     5281 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     5341 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     5401 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     5461 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     5521 gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact attaactggc
     5581 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     5641 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     5701 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     5761 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     5821 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     5881 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     5941 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     6001 gaccccgtag a
//
