LOCUS       sequence-190364-        8045 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8045)
  TITLE     Collier lab plasmids
REFERENCE   2  (bases 1 to 8045)
  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"
COMMENT     Sequence Label: pET-15b LF WT
FEATURES             Location/Qualifiers
     source          1..8045
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     582..601
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(621..698)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     904..923
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1007..1026
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1007..1025
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1026..1050
                     /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             1065..1087
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1106..1123
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1133..1150
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(3553..3571)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3567..3614
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(3789..3794,3795..3811,3812..3817))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(3844..3862)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3930..4034
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4035..4103,4104..4895)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4253..4272)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5066..5654
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5555..5574
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5808..5825
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5840..5979
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5905..5924)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6065..6087
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6081..6272)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg
       61 cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc
      121 gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga
      181 aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc
      241 ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta
      301 caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg
      361 gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg
      421 gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg
      481 taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg
      541 gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg
      601 tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat
      661 gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt
      721 atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc
      781 cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc
      841 caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc
      901 atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc
      961 cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat acgactcact
     1021 ataggggaat tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag
     1081 aaggagatat accatgggca gcagccatca tcatcatcat cacagcagcg gcctggtgcc
     1141 gcgcggcagc catatgctcg aggatccggc gggcggtcat ggtgatgtag gtatgcacgt
     1201 aaaagagaaa gagaaaaata aagatgagaa taagagaaaa gatgaagaac gaaataaaac
     1261 acaggaagag catttaaagg aaatcatgaa acacattgta aaaatagaag taaaagggga
     1321 ggaagctgtt aaaaaagagg cagcagaaaa gctacttgag aaagtaccat ctgatgtttt
     1381 agagatgtat aaagcaattg gaggaaagat atatattgtg gatggtgata ttacaaaaca
     1441 tatatcttta gaagcattat ctgaagataa gaaaaaaata aaagacattt atgggaaaga
     1501 tgctttatta catgaacatt atgtatatgc aaaagaagga tatgaacccg tacttgtaat
     1561 ccaatcttcg gaagattatg tagaaaatac tgaaaaggca ctgaacgttt attatgaaat
     1621 aggtaagata ttatcaaggg atattttaag taaaattaat caaccatatc agaaattttt
     1681 agatgtatta aataccatta aaaatgcatc tgattcagat ggacaagatc ttttatttac
     1741 taatcagctt aaggaacatc ccacagactt ttctgtagaa ttcttggaac aaaatagcaa
     1801 tgaggtacaa gaagtatttg cgaaagcttt tgcatattat atcgagccac agcatcgtga
     1861 tgttttacag ctttatgcac cggaagcttt taattacatg gataaattta acgaacaaga
     1921 aataaatcta tccttggaag aacttaaaga tcaacggatg ctgtcaagat atgaaaaatg
     1981 ggaaaagata aaacagcact atcaacactg gagcgattct ttatctgaag aaggaagagg
     2041 acttttaaaa aagctgcaga ttcctattga gccaaagaaa gatgacataa ttcattcttt
     2101 atctcaagaa gaaaaagagc ttctaaaaag aatacaaatt gatagtagtg attttttatc
     2161 tactgaggaa aaagagtttt taaaaaagct acaaattgat attcgtgatt ctttatctga
     2221 agaagaaaaa gagcttttaa atagaataca ggtggatagt agtaatcctt tatctgaaaa
     2281 agaaaaagag tttttaaaaa agctgaaact tgatattcaa ccatatgata ttaatcaaag
     2341 gttgcaagat acaggagggt taattgatag tccgtcaatt aatcttgatg taagaaagca
     2401 gtataaaagg gatattcaaa atattgatgc tttattacat caatccattg gaagtacctt
     2461 gtacaataaa atttatttgt atgaaaatat gaatatcaat aaccttacag caaccctagg
     2521 tgcggattta gttgattcca ctgataatac taaaattaat agaggtattt tcaatgaatt
     2581 caaaaaaaat ttcaaatata gtatttctag taactatatg attgttgata taaatgaaag
     2641 gcctgcatta gataatgagc gtttgaaatg gagaatccaa ttatcaccag atactcgagc
     2701 aggatattta gaaaatggaa agcttatatt acaaagaaac atcggtctgg aaataaagga
     2761 tgtacaaata attaagcaat ccgaaaaaga atatataagg attgatgcga aagtagtgcc
     2821 aaagagtaaa atagatacaa aaattcaaga agcacagtta aatataaatc aggaatggaa
     2881 taaagcatta gggttaccaa aatatacaaa gcttattaca ttcaacgtgc ataatagata
     2941 tgcatccaat attgtagaaa gtgcttattt aatattgaat gaatggaaaa ataatattca
     3001 aagtgatctt ataaaaaagg taacaaatta cttagttgat ggtaatggaa gatttgtttt
     3061 taccgatatt actctcccta atatagctga acaatataca catcaagatg agatatatga
     3121 gcaagttcat tcaaaagggt tatatgttcc agaatcccgt tctatattac tccatggacc
     3181 ttcaaaaggt gtagaattaa ggaatgatag tgagggtttt atacacgaat ttggacatgc
     3241 tgtggatgat tatgctggat atctattaga taagaaccaa tctgatttag ttacaaattc
     3301 taaaaaattc attgatattt ttaaggaaga agggagtaat ttaacttcgt atgggagaac
     3361 aaatgaagcg gaattttttg cagaagcctt taggttaatg cattctacgg accatgctga
     3421 acgtttaaaa gttcaaaaaa atgctccgaa aactttccaa tttattaacg atcagattaa
     3481 gttcattatt aactcataat gaggatccgg ctgctaacaa agcccgaaag gaagctgagt
     3541 tggctgctgc caccgctgag caataactag cataacccct tggggcctct aaacgggtct
     3601 tgaggggttt tttgctgaaa ggaggaacta tatccggata tcccgcaaga ggcccggcag
     3661 taccggcata accaagccta tgcctacagc atccagggtg acggtgccga ggatgacgat
     3721 gagcgcattg ttagatttca tacacggtgc ctgactgcgt tagcaattta actgtgataa
     3781 actaccgcat taaagcttat cgatgataag ctgtcaaaca tgagaattct tgaagacgaa
     3841 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     3901 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     3961 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     4021 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     4081 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     4141 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     4201 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     4261 gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt
     4321 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     4381 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     4441 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     4501 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     4561 gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac
     4621 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     4681 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     4741 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     4801 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     4861 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     4921 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     4981 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     5041 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     5101 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     5161 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     5221 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     5281 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     5341 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     5401 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     5461 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     5521 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     5581 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     5641 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     5701 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     5761 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     5821 gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     5881 cacaccgcaa tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta
     5941 tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc
     6001 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     6061 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag
     6121 ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc ctgttcatcc
     6181 gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc
     6241 atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg
     6301 ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat
     6361 gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg
     6421 cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt
     6481 gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc
     6541 gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt
     6601 gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat
     6661 tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc
     6721 acgatcatgc gcacccgtgg ccaggaccca acgctgcccg agatgcgccg cgtgcggctg
     6781 ctggagatgg cggacgcgat ggatatgttc tgccaagggt tggtttgcgc attcacagtt
     6841 ctccgcaaga attgattggc tccaattctt ggagtggtga atccgttagc gaggtgccgc
     6901 cggcttccat tcaggtcgag gtggcccggc tccatgcacc gcgacgcaac gcggggaggc
     6961 agacaaggta tagggcggcg cctacaatcc atgccaaccc gttccatgtg ctcgccgagg
     7021 cggcataaat cgccgtgacg atcagcggtc caatgatcga agttaggctg gtaagagccg
     7081 cgagcgatcc ttgaagctgt ccctgatggt cgtcatctac ctgcctggac agcatggcct
     7141 gcaacgcggg catcccgatg ccgccggaag cgagaagaat cataatgggg aaggccatcc
     7201 agcctcgcgt cgcgaacgcc agcaagacgt agcccagcgc gtcggccgcc atgccggcga
     7261 taatggcctg cttctcgccg aaacgtttgg tggcgggacc agtgacgaag gcttgagcga
     7321 gggcgtgcaa gattccgaat accgcaagcg acaggccgat catcgtcgcg ctccagcgaa
     7381 agcggtcctc gccgaaaatg acccagagcg ctgccggcac ctgtcctacg agttgcatga
     7441 taaagaagac agtcataagt gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc
     7501 tgactgggtt gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct
     7561 aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac tgtcgtgcca
     7621 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg
     7681 tggtttttct tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct
     7741 gagagagttg cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga
     7801 tggtggttaa cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg
     7861 agatatccgc accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca
     7921 tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg
     7981 tttgttgaaa accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt
     8041 gattg
//
