LOCUS       sequence-392337-        8155 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8155)
  TITLE     George Cross Lab Plasmids
REFERENCE   2  (bases 1 to 8155)
  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"
COMMENT     Sequence Label: pLEW100v5-177-HYG
FEATURES             Location/Qualifiers
     source          1..8155
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..19
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    22..40
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             236..1888
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKRLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(302..320)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     promoter        complement(2772..2790)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2773..2790)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3080..3099)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(3309..3328)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3488..3506)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3574..3678
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3679..3747,3748..4539)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHSMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3897..3916)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4710..5298
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5199..5218
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5452..5469
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(6149..6196)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      complement(6281..6328)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(6691..7716)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(6691..7716)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     primer_bind     complement(7860..7879)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7861..7879)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 tccctatcag tgatagagat ctccctatca gtgatagaga tccctgagta ctgagtttaa
       61 catgttctcg tcccgggctg cacgcgcctt cgagtttttt ttccttttcc ccattttttt
      121 caacttgaag acttcaatta caccaaaaag taaaattcac aagcttggaa ttcctttgtg
      181 ttacattctt gaatgtcgct cgcagtgaca ttagcattcc ggtactgttg gtaaaatgga
      241 agacgccaaa aacataaaga aaggcccggc gccattctat cctctagagg atggaaccgc
      301 tggagagcaa ctgcataagg ctatgaagag atacgccctg gttcctggaa caattgcttt
      361 tacagatgca catatcgagg tgaacatcac gtacgcggaa tacttcgaaa tgtccgttcg
      421 gttggcagaa gctatgaaac gatatgggct gaatacaaat cacagaatcg tcgtatgcag
      481 tgaaaactct cttcaattct ttatgccggt gttgggcgcg ttatttatcg gagttgcagt
      541 tgcgcccgcg aacgacattt ataatgaacg tgaattgctc aacagtatga acatttcgca
      601 gcctaccgta gtgtttgttt ccaaaaaggg gttgcaaaaa attttgaacg tgcaaaaaaa
      661 attaccaata atccagaaaa ttattatcat ggattctaaa acggattacc agggatttca
      721 gtcgatgtac acgttcgtca catctcatct acctcccggt tttaatgaat acgattttgt
      781 accagagtcc tttgatcgtg acaaaacaat tgcactgata atgaattcct ctggatctac
      841 tgggttacct aagggtgtgg cccttccgca tagaactgcc tgcgtcagat tctcgcatgc
      901 cagagatcct atttttggca atcaaatcat tccggatact gcgattttaa gtgttgttcc
      961 attccatcac ggttttggaa tgtttactac actcggatat ttgatatgtg gatttcgagt
     1021 cgtcttaatg tatagatttg aagaagagct gtttttacga tcccttcagg attacaaaat
     1081 tcaaagtgcg ttgctagtac caaccctatt ttcattcttc gccaaaagca ctctgattga
     1141 caaatacgat ttatctaatt tacacgaaat tgcttctggg ggcgcacctc tttcgaaaga
     1201 agtcggggaa gcggttgcaa aacgcttcca tcttccaggg atacgacaag gatatgggct
     1261 cactgagact acatcagcta ttctgattac acccgagggg gatgataaac cgggcgcggt
     1321 cggtaaagtt gttccatttt ttgaagcgaa ggttgtggat ctggataccg ggaaaacgct
     1381 gggcgttaat cagagaggcg aattatgtgt cagaggacct atgattatgt ccggttatgt
     1441 aaacaatccg gaagcgacca acgccttgat tgacaaggat ggatggctac attctggaga
     1501 catagcttac tgggacgaag acgaacactt cttcatagtt gaccgcttga agtctttaat
     1561 taaatacaaa ggatatcagg tggcccccgc tgaattggaa tcgatattgt tacaacaccc
     1621 caacatcttc gacgcgggcg tggcaggtct tcccgacgat gacgccggtg aacttcccgc
     1681 cgccgttgtt gttttggagc acggaaagac gatgacggaa aaagagatcg tggattacgt
     1741 cgccagtcaa gtaacaaccg cgaaaaggtt gcgcggagga gttgtgtttg tggacgaagt
     1801 accgaaaggt cttaccggaa aactcgacgc aagaaaaatc agagagatcc tcataaaggc
     1861 caagaagggc ggaaagtcca aattgtaaaa tgtaactgta ttcagcgatg acgaaattct
     1921 tagctattgt aatattatat gcaaattgat gaatggtaat tttgtaattg tgggtcactg
     1981 tactatttta acgaataata aaatcaggta taggtaacta aaaaggaatt cgagcttgga
     2041 tcctgcccat ttagttggct tttcccttgt ctcgtgtctt ttccgtggaa aggttcccgg
     2101 agtaatctga tggcacagca gggaggtgcg cctgcaggtt ggttaggaag gggggatgat
     2161 gtaaaagaag aaaatggggg gatatcttat gtttaagagg ataaataatg tgaaggggct
     2221 ttatatgctt gcttggttgt ctcgttggtg catggggaat ctgcatgttt gctttggagc
     2281 acgcgtggta catcaatggg atcatatctt gctgcctccc gcagtcaccg tgcggagctg
     2341 ccggtgcccc gtttctcata ggcgtgtgca ccttccgtct gagacctgta aatggttaaa
     2401 aaggaatata taattacctt ttgaaaagtg gtaaaaaacg aatatatgtt tttttttggt
     2461 tttaatacgc tcttttgtgt gtatgagagg aataaataag tgtgtgtgtg tgtgtgtgtt
     2521 gttgttgttg aaaaaatgaa ggacacgaaa gcgctcatta ggggtgggga tacaccaggc
     2581 ataattatcc gaaaatattt tggtgcacca aataagtgaa catactgacg aaatcaaaga
     2641 gttggaggat ggatagggag gcctcattgg cagtggtaaa ttgatttgac ttgaatatga
     2701 taccctctta ctgtttcatt gccactacaa gttggtttcc ttcccctgca ggcatgcaag
     2761 ctagcttgta ttctatagtg tcacctaaat cgtatgtgta tgatacataa ggttatgtat
     2821 taattgtagc cgcgttctaa cgacaatatg tacaagccta attgtgtagc atctggctta
     2881 ctgaagcaga ccctatcatc tctctcgtaa actgccgtca gagtcggttt ggttggacga
     2941 accttctgag tttctggtaa cgccgtcccg cacccggaaa tggtcagcga accaatcagc
     3001 agggtcatcg ctagccagat cctctacgcc ggacgcatcg tggccggcat caccggcgcc
     3061 acaggtgcgg ttgctggcgc ctatatcgcc gacatcaccg atggggaaga tcgggctcgc
     3121 cacttcgggc tcatgagcgc ttgtttcggc gtgggtatgg tggcaggccc cgtggccggg
     3181 ggactgttgg gcgccatctc cttgcaccat tccttgcggc ggcggtgctc aacggcctca
     3241 acctactact gggctgcttc ctaatgcagg agtcgcataa gggagagcgt cgaatggtgc
     3301 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     3361 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     3421 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     3481 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     3541 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     3601 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     3661 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     3721 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     3781 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     3841 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gtacttttaa agttctgcta
     3901 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     3961 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     4021 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     4081 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca cagcatgggg
     4141 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     4201 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     4261 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     4321 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     4381 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     4441 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     4501 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     4561 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     4621 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     4681 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     4741 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     4801 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     4861 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     4921 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     4981 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     5041 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     5101 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     5161 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     5221 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     5281 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     5341 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     5401 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     5461 gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     5521 attcattaat gcagaattcg agctctgcag gaattcgtgc tctttaatgg tccttatacg
     5581 aacgaatatt aaacaatgcg cagttaacgc tattatacac aataactttt aatgtctgcc
     5641 atattaatta caagtgtgca acattaaata caagtgtgta acgttaattt gcaagtttga
     5701 aacaatgttc tttagtgttt aatgtgtgca acaaagctat ttaatggtcc ttatacgaac
     5761 gaatattaaa caatgcgcag ttaacgctat tatacacaat aacttttaat gtgtgccata
     5821 ttaattacgg atcgcggccg cgatccgtgt gcaacattaa atacaagtgt gtaacattaa
     5881 tttgcaagtt tgcaacaatg ttctttagtg tttaatgggt gcaacaaagc tatttaatgg
     5941 tccttatacg aacgaatatt aaacaatccg cagttaacgc tattatacac aataactttt
     6001 aatgtgtccc atattaatta caagtgtgca acattaaata caagtgtgta acattaattt
     6061 gcaagtttgc aacaatgttc tttagtgttt aatgggtacc gaattcctgc aggggcttaa
     6121 gcctatccgg atatagttct cctttcagca aaaaacccct caagacccgt ttagaggccc
     6181 caaggggtta tgctagttat tgctcagcgg tggcagcagc caactcagct tcctttcggg
     6241 ctttgttagc agccctatcc ggatatagtt ctcctttcag caaaaaaccc ctcaagaccc
     6301 gtttagaggc cccaaggggt tatgctagtt attgctcagc ggtggcagca gccaactcag
     6361 cttcctttcg ggctttgtta gcagcattta aatgcagaat actgcataga taacaaacgc
     6421 atcaaacaca actggtggca ctatttcaat catgtcgaca caccaagacg agaaaatttg
     6481 agttaaacaa gatgcctaca accctatatt tattatcttc aagtgatctt ctcttctctt
     6541 gtgcgctgta cgtaaatgtg ttgcaaataa ggcatagggc tgagtacagg caccacacgc
     6601 gaggaaaatc ataagcacta gccgtgtcaa ctcacagcga ctacagaaca attttggcca
     6661 cacaacccgg tgttaggatc tccgaggcct ctattccttt gccctcggac gagtgctggg
     6721 gcgtcggttt ccactatcgg cgagtacttc tacacagcca tcggtccaga cggccgcgct
     6781 tctgcgggcg atttgtgtac gcccgacagt cccggctccg gatcggacga ttgcgtcgca
     6841 tcgaccctgc gcccaagctg catcatcgaa attgccgtca accaagctct gatagagttg
     6901 gtcaagacca atgcggagca tatacgcccg gagccgcggc gatcctgcaa gctccggatg
     6961 cctccgctcg aagtagcgcg tctgctgctc catacaagcc aaccacggcc tccagaagaa
     7021 gatgttggcg acctcgtatt gggaatcccc gaacatcgcc tcgctccagt caatgaccgc
     7081 tgttatgcgg ccattgtccg tcaggacatt gttggagccg aaatccgcgt gcacgaggtg
     7141 ccggacttcg gggcagtcct cggcccaaag catcagctca tcgagagcct gcgcgacgga
     7201 cgcactgacg gtgtcgtcca tcacagtttg ccagtgatac acatggggat cagcaatcgc
     7261 gcatatgaaa tcacgccatg tagtgtattg accgattcct tgcggtccga atgggccgaa
     7321 cccgctcgtc tggctaagat cggccgcagc gatcgcatcc atggcctccg cgaccggctg
     7381 cagaacagcg ggcagttcgg tttcaggcag gtcttgcaac gtgacaccct gtgcacggcg
     7441 ggagatgcaa taggtcaggc tctcgctgaa ttccccaatg tcaagcactt ccggaatcgg
     7501 gagcgcggcc gatgcaaagt gccgataaac ataacgatct ttgtagaaac catcggcgca
     7561 gctatttacc cgcaggacat atccacgccc tcctacatcg aagctgaaag cacgagattc
     7621 ttcgccctcc gagagctgca tcaggtcgga gacgctgtcg aacttttcga tcagaaactt
     7681 ctcgacagac gtcgcggtga gttcaggctt tttcatcact agttctagag cttattttat
     7741 ggcagcaacg agaccttacg taaagcaaaa ctaaataaga gcggagactg caatgcagag
     7801 taaaaaaaaa aaaacatgaa tttcagaaga ccttgctgtg cccccggtac gggagatctc
     7861 cctatagtga gtcgtattaa tcaggtacct agctttccac ccagcgcggg tgcattctgg
     7921 ctcttatata tacttattgt catgacagag tatattgtac tgtgttgata agggacgggt
     7981 aactgtattg aagagccgat gcttttgaca tgttagatat aatatgtttt attgtaaagt
     8041 caatacaaca cacaatagga taataatgat aaagttaaaa aagtatatat agtaatagaa
     8101 atatatctta tataggaaag attaagcagt aaaagtagcg cttacggcgt acggt
//
