LOCUS       sequence-422154-        6258 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses outer membrane protein F.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6258)
  AUTHORS   Hiller
  TITLE     Hiller Lab Plasmids 2023
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6258)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: OmpF
FEATURES             Location/Qualifiers
     source          1..6258
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1199..1221
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1236..1260
                     /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)."
     primer_bind     complement(1260..1279)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1261..1279)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1363..1382)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1588..1665
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1666..2748
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(1685..1704)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3557..3748
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3742..3764)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3850..3989
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3905..3924
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4004..4021)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4175..4763)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4255..4274)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4874..4893)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4885..5700
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4885..5700
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4958..4977)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5792..6247)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5929..5950)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6141..6160
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagttagaac tggtaaacga
      181 tacccacagc aacggtgtcg tctgaaccta cgcccagttt gttgtcagaa tcgatctggt
      241 tgatgatgta gtcaacatag gtggacatgt ttttgttgaa gtagtaggtt gcgcccactt
      301 caaagtagtt caccagatca acatcaccga taccttctac gtctttcgct ttagatttgg
      361 tgtaagcgat ggacggacgc agaccgaaat cgaactggta ttgcgcaact aacagaacgt
      421 cttgcgtttt gttggcgaag ccgctggtgt ttgtaaattt attagtgatc ggcgtagcgt
      481 tacgggtttc accgtagttc gctgccaggt agatgttgtt cgcgtcgtac ttcagaccag
      541 tagcccactg ttcagctttt ttaccgttgc caagaggttg agcttcttgc aggttggtac
      601 ggtcagctgc accataagca ccaacgatac caaagccttc gtattcgtag ctgatagaac
      661 cgccaacacc gtcgccgtta gaacggcgtg cagtgtcacg ctcgttttta cccaggtact
      721 gaacagcgaa gttcaggcca tcaaccagac caaagaagtt ggagttacga taggtagcaa
      781 cgccgccaac acgaccaacg aagaagtcat cgctgtatgc agtatcacca ccaaattctg
      841 gcagcatatc ggtgtaaccc agtgcatcat aaaccacacc gtagttacgg ccgtaatcga
      901 aagaaccaac gtcagcgtat ttaagacccg cgaatgccag acgcgttttg ttaccagttt
      961 gagcgtcagc gccttcagag ttgttaccct ggaagttata ttcccactga ccataaccgg
     1021 tcagatcgga attgatttga gtttcccctt taaaaccaag acgggcatag gtcatgtcgc
     1081 cattgccacc gtaactgttt tcaccgttac ccttggaaaa ataatgcaga ccaacagctt
     1141 taccgtacag atctactttg ttgccatctt tgttatagat ttctgcagcc atggtatatc
     1201 tccttcttaa agttaaacaa aattatttct agaggggaat tgttatccgc tcacaattcc
     1261 cctatagtga gtcgtattaa tttcgcggga tcgagatctc gatcctctac gccggacgca
     1321 tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca
     1381 ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta
     1441 tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc
     1501 ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt
     1561 cgcataaggg agagcgtcga gatcccggac accatcgaat ggcgcaaaac ctttcgcggt
     1621 atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg
     1681 ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac
     1741 caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg
     1801 aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc
     1861 gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct
     1921 cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa
     1981 gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac
     2041 tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg
     2101 ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac
     2161 ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta
     2221 gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc
     2281 actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt
     2341 tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc
     2401 aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt
     2461 gcggatatct cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg
     2521 ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg
     2581 caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa
     2641 agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     2701 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     2761 taatgtaagt tagctcactc attaggcacc gggatctcga ccgatgccct tgagagcctt
     2821 caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac
     2881 tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctctggg tcattttcgg
     2941 cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat
     3001 cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa
     3061 gcaggccatt atcgccggca tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt
     3121 gaggacccgg ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg
     3181 cgagcgaacg tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat
     3241 ggtcttcggt ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat
     3301 tatgttccgg atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt
     3361 attaacgaag cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac
     3421 cgccagttgt ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg
     3481 tatcgtgagc atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc
     3541 ttacacggag gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat
     3601 cagaagccag acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc
     3661 agacatctgt gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg
     3721 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     3781 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     3841 gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac
     3901 tatgcggcat cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca
     3961 gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc
     4021 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4081 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4141 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     4201 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4261 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4321 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4381 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4441 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4501 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4561 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     4621 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4681 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4741 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4801 agtggaacga aaactcacgt taagggattt tggtcatgaa caataaaact gtctgcttac
     4861 ataaacagta atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctctagg
     4921 ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat
     4981 gtcgggcaat caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg
     5041 tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta
     5101 aactggctga cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat
     5161 gatgcatggt tactcaccac tgcgatcccc gggaaaacag cattccaggt attagaagaa
     5221 tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat
     5281 tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg
     5341 caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc
     5401 tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca
     5461 gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata
     5521 ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta
     5581 tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt
     5641 attgataatc ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa
     5701 gaattaattc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     5761 tccgcgcaca tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa
     5821 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     5881 atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac
     5941 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     6001 ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     6061 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     6121 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     6181 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     6241 ggcgcgtccc attcgcca
//
