LOCUS       sequence-204358-        6029 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  plasmid for expression of computationally designed colicin 
            endonuclease and immunity pair, des16.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6029)
  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 6029)
  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-colEdes16/Imdes16
FEATURES             Location/Qualifiers
     source          1..6029
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      16..604
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     505..524
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     758..775
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    790..929
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(855..874)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1015..1037
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1031..1222)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     3074..3093
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3113..3190)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3396..3415
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3499..3518
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3499..3517
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3518..3542
                     /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             3557..3579
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4260..4277
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4330..4348)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4344..4391
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4428..4883
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4515..4534)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4725..4746
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4909..5013
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5014..5082,5083..5874)
                     /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(5232..5251)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
       61 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
      121 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
      181 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
      241 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
      301 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
      361 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
      421 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
      481 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
      541 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
      601 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
      661 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
      721 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
      781 ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa
      841 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc
      901 tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc
      961 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     1021 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa
     1081 gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct
     1141 cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg
     1201 cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg
     1261 taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg
     1321 cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga
     1381 gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg
     1441 gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc
     1501 gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg
     1561 cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct
     1621 aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc
     1681 gcacccgtgg ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg
     1741 cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca
     1801 ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg
     1861 ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag
     1921 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc
     1981 gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg
     2041 ctttccagtc gggaaactgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     2101 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca
     2161 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt
     2221 gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt
     2281 cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg
     2341 taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa
     2401 cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc
     2461 cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca
     2521 gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac
     2581 ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac
     2641 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag
     2701 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc
     2761 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca
     2821 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt
     2881 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc
     2941 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca
     3001 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct
     3061 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca
     3121 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt
     3181 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc
     3241 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg
     3301 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
     3361 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
     3421 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
     3481 atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
     3541 cccctctaga aataattttg tttaacttta agaaggagat ataccatgga gtccaaacgt
     3601 aataaaccgg gtaaagcgac cggtaaaggt aaaccagttg gtgataaatg gctggacgac
     3661 gcgggcaaag actccggtgc gccgatcccg gatcgtatcg cggacaagct gcgtgacaag
     3721 gaattcaaaa acttcgacga cttcaagaaa aagttctgga aagaagtttc taaggacccg
     3781 gacctgtcta aacagttcaa aggttctaac aaactgtcta tctctcaggg tcaggcgccg
     3841 ttcgcgcgtc agaaggacca ggttggtggt cgtcgccgtt tcgaactgca ccacgacaaa
     3901 ccgatttctc aggacggtgg tgtttacgac atgaacaaca tccgtgttac caccccgaaa
     3961 cgtcacatcg acatccaccg gggtaagtaa aaatggaact gaaacactct atttctgact
     4021 acaccgaagc ggagttcctg gaattcgtta aaaagtcttg cgaggcggaa ctggaaggtc
     4081 gtgaggaagt cgctaagcgt ctcgttgaag aattcgaacg tctgaccgaa cacccggatg
     4141 gttctgacct gatctactac ccgcgtgacg accgcgaaga ctctccggaa ggtattgtta
     4201 aggagattaa agaatggcgt gcggcgaatg gtaaatctgg tttcaaacag ggcctcgagc
     4261 accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     4321 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     4381 ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg cgccctgtag
     4441 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     4501 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     4561 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     4621 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     4681 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4741 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     4801 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     4861 caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct
     4921 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     4981 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     5041 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     5101 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     5161 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     5221 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     5281 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     5341 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     5401 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     5461 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     5521 gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac aacgttgcgc
     5581 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     5641 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     5701 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     5761 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     5821 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     5881 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     5941 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     6001 ttccactgag cgtcagaccc cgtagaaaa
//
