LOCUS       sequence-204348-        6020 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  plasmid for expression of computationally designed colicin 
            endonuclease and immunity pair, des9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6020)
  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 6020)
  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-colEdes9/Imdes9
FEATURES             Location/Qualifiers
     source          1..6020
                     /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     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             4251..4268
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4321..4339)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4335..4382
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4419..4874
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4506..4525)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4716..4737
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4900..5004
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5005..5073,5074..5865)
                     /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(5223..5242)
                     /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 tgtctgcgat ataggcgcca
     3421 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
     3481 atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
     3541 cccctctaga aataattttg tttaacttta agaaggagat ataccatgga gagtaaacgg
     3601 aataagccag ggaaggcgac aggtaaaggt aaaccagttg gtgataaatg gctggatgat
     3661 gcaggtaaag attcaggagc gccaattcca gatcgcattg ctgataagtt gcgtgataaa
     3721 gaatttaaaa actttgacga tttccggaag aaattctggg aagaagtgtc aaaagatccc
     3781 gatcttagta agcaatttaa aggcgcgaat caagaaaaca ttaagcaagg acaagcacct
     3841 tttgcaaggc aaaaagacca agtaggtggt aggagggtct ttgaattaca tcatgataaa
     3901 ccaatcagtc aggatggtgg tgtctatgat atgaataata tcagagtgac cacacctaag
     3961 cgacatattg atattcatcg gggtaagtaa aaatggaact gaaacatagt attagtgatt
     4021 ataccgaggc tgaatttctg gagtttgtaa aaaaaatatt agagttaact gacgaacaga
     4081 aacaggctca gctggtggac gagtttgagc gattaactga gcacccagac ggttcagatc
     4141 tgatttatta tcctcgcgat gacagggaag atagtcctga agggattgtc aaggaaatta
     4201 aagaatggcg agctgctaac ggtaagtcag gatttaaaca gggcctcgag caccaccacc
     4261 accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca
     4321 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     4381 tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta gcggcgcatt
     4441 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     4501 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     4561 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     4621 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     4681 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     4741 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     4801 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     4861 aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     4921 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     4981 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     5041 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     5101 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     5161 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     5221 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     5281 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     5341 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5401 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5461 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5521 aacgacgagc gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta
     5581 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5641 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5701 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     5761 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     5821 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     5881 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     5941 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6001 gcgtcagacc ccgtagaaaa
//
