LOCUS       sequence-11278-e        9261 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 9261)
  TITLE     George Cross Lab Plasmids
REFERENCE   2  (bases 1 to 9261)
  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: pLEW13
FEATURES             Location/Qualifiers
     source          1..9261
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1278..1298
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4388..4407)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4998..5017
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             5572..6195
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(5645..5664)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(6497..6515)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(6498..6515)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(6806..6825)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(7036..7055)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7155..7177
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7215..7233)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7301..7405
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7406..7474,7475..8266)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7624..7643)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8437..9025
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8926..8945
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9179..9196
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ctgaccgtat catgatgact ttctccatca tcccatcccc caaggtgtcc gacactgtcg
       61 tcgagccgta caatacgact ctctccgtgc accaacttgt ggaaaactcc gatgagtcga
      121 tgtgcattga caacgaggca ctgtacgata tttgcttccg caccctgaaa ctgacaacac
      181 caacgttcgg tgacctgaac cacttggtgt ctgctgttgt gtccggcgtc acctgctgcc
      241 tgcgcttccc tggtcagttg aactctgacc tccgtaagtt ggctgtgaac cttgtcccat
      301 tcccgcgtct gcacttcttc atgatgggct tcgccccgct gaccagccgc ggctcgcagc
      361 agtaccgcgg tctctccgtg cccgagctaa cgcagcagat gttcgatgcg aaaaacatga
      421 tgcaagctgc agatcctcgt cacggccgct acctgacagc gtctgcactc ttccgcggcc
      481 gcatgtcgac gaaggaggtt gatgagcaga tgctgaacgt gcagaacaag aactcgtcct
      541 acttcattga gtggatcgat cccgaacaac atcaagtcct ctgtttgcga tatcccaccc
      601 aagggactca agatggctgt caccttcatt ggcaacaaca cctgcatcca ggagatgttc
      661 cgccgtgtgg gagagcagtt caccctcatg ttccgtcgca aggcgttctt gcactggtac
      721 actggcgagg gtatggacga gatggaattc acggaggcag agtccaacat gaacgatctc
      781 gtgtctgagt accagcagta ccaggatgcc acgattgagg aggagggcga gttcgacgag
      841 gaggagcaat actagacgcg gacggggcat ttcccgttcg tcattagcag taggtaatga
      901 agatgctcga gggtgctcaa gctgtgtagc gcacgcgttt ccttacatat ttctctaaca
      961 ggcacggaag cctaacaaat acacttggct tatttttttg ccccctcatg tcttgtacaa
     1021 atatttgcga tagcttagct atcagccaca ttaatcaaac aagtatacca acaagcccga
     1081 aaacataaac tcaactgcaa cgaagctggg ctgcacgcgc cttcgagttt tttttccttt
     1141 tccccatttt tttcaacttg aagacttcaa ttacaccaaa aagtaaaatt cacaagctga
     1201 tctgatccgg atttactaac tggaagaggc actaaatgaa cacgattaac atcgctaaga
     1261 acgaattcct cgagcctcca aaaaagaaga gaaaggtcga attctctgac atcgaactgg
     1321 ctgctatccc gttcaacact ctggctgacc attacggtga gcgtttagct cgcgaacagt
     1381 tggcccttga gcatgagtct tacgagatgg gtgaagcacg cttccgcaag atgtttgagc
     1441 gtcaacttaa agctggtgag gttgcggata acgctgccgc caagcctctc atcactaccc
     1501 tactccctaa gatgattgca cgcatcaacg actggtttga ggaagtgaaa gctaagcgcg
     1561 gcaagcgccc gacagccttc cagttcctgc aagaaatcaa gccggaagcc gtagcgtaca
     1621 tcaccattaa gaccactctg gcttgcctaa ccagtgctga caatacaacc gttcaggctg
     1681 tagcaagcgc aatcggtcgg gccattgagg acgaggctcg cttcggtcgt atccgtgacc
     1741 ttgaagctaa gcacttcaag aaaaacgttg aggaacaact caacaagcgc gtagggcacg
     1801 tctacaagaa agcatttatg caagttgtcg aggctgacat gctctctaag ggtctactcg
     1861 gtggcgaggc gtggtcttcg tggcataagg aagactctat tcatgtagga gtacgctgca
     1921 tcgagatgct cattgagtca accggaatgg ttagcttaca ccgccaaaat gctggcgtag
     1981 taggtcaaga ctctgagact atcgaactcg cacctgaata cgctgaggct atcgcaaccc
     2041 gtgcaggtgc gctggctggc atctctccga tgttccaacc ttgcgtagtt cctcctaagc
     2101 cgtggactgg cattactggt ggtggctatt gggctaacgg tcgtcgtcct ctggcgctgg
     2161 tgcgtactca cagtaagaaa gcactgatgc gctacgaaga cgtttacatg cctgaggtgt
     2221 acaaagcgat taacattgcg caaaacaccg catggaaaat caacaagaaa gtcctagcgg
     2281 tcgccaacgt aatcaccaag tggaagcatt gtccggtcga ggacatccct gcgattgagc
     2341 gtgaagaact cccgatgaaa ccggaagaca tcgacatgaa tcctgaggct ctcaccgcgt
     2401 ggaaacgtgc tgccgctgct gtgtaccgca aggacaaggc tcgcaagtct cgccgtatca
     2461 gccttgagtt catgcttgag caagccaata agtttgctaa ccataaggcc atctggttcc
     2521 cttacaacat ggactggcgc ggtcgtgttt acgctgtgtc aatgttcaac ccgcaaggta
     2581 acgatatgac caaaggactg cttacgctgg cgaaaggtaa accaatcggt aaggaaggtt
     2641 actactggct gaaaatccac ggtgcaaact gtgcgggtgt cgataaggtt ccgttccctg
     2701 agcgcatcaa gttcattgag gaaaaccacg agaacatcat ggcttgcgct aagtctccac
     2761 tggagaacac ttggtgggct gagcaagatt ctccgttctg cttccttgcg ttctgctttg
     2821 agtacgctgg ggtacagcac cacggcctga gctataactg ctcccttccg ctggcgtttg
     2881 acgggtcttg ctctggcatc cagcacttct ccgcgatgct ccgagatgag gtaggtggtc
     2941 gcgcggttaa cttgcttcct agtgaaaccg ttcaggacat ctacgggatt gttgctaaga
     3001 aagtcaacga gattctacaa gcagacgcaa tcaatgggac cgataacgaa gtagttaccg
     3061 tgaccgatga gaacactggt gaaatctctg agaaagtcaa gctgggcact aaggcactgg
     3121 ctggtcaatg gctggcttac ggtgttactc gcagtgtgac taagcgttca gtcatgacgc
     3181 tggcttacgg gtccaaagag ttcggcttcc gtcaacaagt gctggaagat accattcagc
     3241 cagctattga ttccggcaag ggtctgatgt tcactcagcc gaatcaggct gctggataca
     3301 tggctaagct gatttgggaa tctgtgagcg tgacggtggt agctgcggtt gaagcaatga
     3361 actggcttaa gtctgctgct aagctgctgg ctgctgaggt caaagataag aagactggag
     3421 agattcttcg caagcgttgc gctgtgcatt gggtaactcc tgatggtttc cctgtgtggc
     3481 aggaatacaa gaagcctatt cagacgcgct tgaacctgat gttcctcggt cagttccgct
     3541 tacagcctac cattaacacc aacaaagata gcgagattga tgcacacaaa caggagtctg
     3601 gtatcgctcc taactttgta cacagccaag acggtagcca ccttcgtaag actgtagtgt
     3661 gggcacacga gaagtacgga atcgaatctt ttgcactgat tcacgactcc ttcggtacca
     3721 ttccggctga cgctgcgaac ctgttcaaag cagtgcgcga aactatggtt gacacatatg
     3781 agtcttgtga tgtactggct gatttctacg accagttcgc tgaccagttg cacgagtctc
     3841 aattggacaa aatgccagca cttccggcta aaggtaactt gaacctccgt gacatcttag
     3901 agtcggactt cgcgttcgcg taacgccaaa tcaatacgac tccggatcct gcccatttgg
     3961 cttttccctt gtctcgtgtc ttttccgtgg aaaggttccc ggagtaatct gatggcacag
     4021 cagggaggtg cgcctgcagg ttggttagga aggggggatg atgtaaaaga agaaaatggg
     4081 gggattcgag cccgggcaca gcaaggtctt ctgaaattca tgtttttttt ttttttactc
     4141 tgcattgcag tctccgctct tatttagttt tgctttacgt aaggtctcgt tgctgccata
     4201 aaataagctt agaggagaaa gaatagtaac cctttcatca aagaaaatag ttcaaacgaa
     4261 tatgcgcgaa atcgtctgcg ttcaggctgg ccaatgcggt aaccagatcg gctcaaagtt
     4321 ctgggaggtg atccggccaa gcttggatgg attgcacgca ggttctccgg ccgcttgggt
     4381 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     4441 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     4501 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     4561 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     4621 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     4681 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     4741 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     4801 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     4861 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     4921 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     4981 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     5041 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     5101 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcggatc ctaacaccgg
     5161 gttgtgtggc caaaattgtt ctgtagtcgc tgtgagttga cacggctagt gcttatgatt
     5221 ttcctcgcgt gtggtgcctg tactcagccc tatgccttat ttgcaacaca tttacgtaca
     5281 gcgcacaaga gaagagaaga tcacttgaag ataataaata tagggttgta ggcatcttgt
     5341 ttaactcaaa ttttctcgtc ttggtgtgtc gacatgattg aaatagtgcc accagttgtg
     5401 tttgatgcgt ttgttatcta tgcagtattc tgcaaggggg cacagcaagg tcttctgaaa
     5461 ttcatgtttt tttttttttt actctgcatt gcagtctccg ctcttattta gttttgcttt
     5521 acgtaaggtc tcgttgctgc cataaaataa gcctagaact agagctttca tatgtctaga
     5581 ttagataaaa gtaaagtgat taacagcgca ttagagctgc ttaatgaggt cggaatcgaa
     5641 ggtttaacaa cccgtaaact cgcccagaag ctaggtgtag agcagcctac attgtattgg
     5701 catgtaaaaa ataagcgggc tttgctcgac gccttagcca ttgagatgtt agataggcac
     5761 catactcact tttgcccttt agaaggggaa agctggcaag attttttacg taataacgct
     5821 aaaagtttta gatgtgcttt actaagtcat cgcgatggag caaaagtaca tttaggtaca
     5881 cggcctacag aaaaacagta tgaaactctc gaaaatcaat tagccttttt atgccaacaa
     5941 ggtttttcac tagagaatgc attatatgca ctcagcgctg tggggcattt tactttaggt
     6001 tgcgtattgg aagatcaaga gcatcaagtc gctaaagaag aaagggaaac acctactact
     6061 gatagtatgc cgccattatt acgacaagct atcgaattat ttgatcacca aggtgcagag
     6121 ccagccttct tattcggcct tgaattgatc atatgcggat tagaaaaaca acttaaatgt
     6181 gaaagtgggt cttaaggatc ccaaaattgt tctgtagtcg ctgtgagttg acacggctag
     6241 tgcttatgat tttcctcgcg tgtggtgcct gtactcagcc ctatgcctta tttgcaacac
     6301 atttacgtac agcgcacaag agaagagaag atcacttgaa gataataaat atagggttgt
     6361 aggcatcttg tttaactcaa attttctcgt cttggtgtgt cgacatgatt gaaatagtgc
     6421 caccagttgt gtttgatgcg tttgttatct atgcagtatt ctgcaaggcc ttgcaggcat
     6481 gcaagctagc ttgtattcta tagtgtcacc taaatcgtat gtgtatgata cataaggtta
     6541 tgtattaatt gtagccgcgt tctaacgaca atatgtacaa gcctaattgt gtagcatctg
     6601 gcttactgaa gcagacccta tcatctctct cgtaaactgc cgtcagagtc ggtttggttg
     6661 gacgaacctt ctgagtttct ggtaacgccg ttccgcaccc cggaaatggt cagcgaacca
     6721 atcagcaggg tcatcgctag ccagatcctc tacgccggac gcatcgtggc cggcatcacc
     6781 ggcgccacag gtgcggttgc tggcgcctat atcgccgaca tcaccgatgg ggaagatcgg
     6841 gctcgccact tcgggctcat gagcgcttgt ttcggcgtgg gtatggtggc aggccccgtg
     6901 gccgggggac tgttgggcgc catctccttg caccattcct tgcggcggcg gtgctcaacg
     6961 gcctcaacct actactgggc tgcttcctaa tgcaggagtc gcataaggga gagcgtcgat
     7021 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     7081 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     7141 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     7201 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     7261 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     7321 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     7381 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     7441 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     7501 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     7561 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     7621 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     7681 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     7741 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     7801 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     7861 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     7921 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     7981 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     8041 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     8101 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     8161 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     8221 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     8281 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     8341 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     8401 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     8461 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     8521 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     8581 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     8641 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     8701 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     8761 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     8821 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     8881 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     8941 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     9001 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     9061 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     9121 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     9181 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     9241 ttggccgatt cattaatgca g
//
