LOCUS       sequence-146182-        6823 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Creating LexA::GADfluw lines from MiMIC lines with inserts into 
            coding introns in Phase 2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6823)
  AUTHORS   Diao F, Ironfield H, Luan H, Diao F, Shropshire WC, Ewer J, Marr E, 
            Potter CJ, Landgraf M, White BH
  TITLE     Plug-and-play genetic access to drosophila cell types using 
            exchangeable exon cassettes.
  JOURNAL   Cell Rep. 2015 Mar 3;10(8):1410-21. doi: 
            10.1016/j.celrep.2015.01.059. Epub 2015 Feb 26.
  PUBMED    25732830
REFERENCE   2  (bases 1 to 6823)
  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"; doi: 
            "10.1016/j.celrep.2015.01.059"; journalName: "Cell Rep"; date: 
            "2015-03-3- 3"; volume: "10"; issue: "8"; pages: "1410-21"
COMMENT     Sequence Label: pBS-KS-attB2-SA(2)-T2A-LexA::GADfluw-Hsp70
FEATURES             Location/Qualifiers
     source          1..6823
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(83..100)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(254..842)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(334..353)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1013..1804,1805..1873))
                     /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     1636..1655
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1874..1978)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2004..2459)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2141..2162)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2353..2372
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2586..2608
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2600..2617
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2601..2617
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2627..2646
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2627..2645
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2681..2750
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     CDS             2918..2971
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2978..2998
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(3008..3259,3260..3613)
                     /codon_start=1
                     /gene="lexA"
                     /product="E. coli DNA-binding protein that represses genes 
                     involved in the SOS response to DNA damage"
                     /label=LexA
                     /note="The N-terminal DNA-binding domain recognizes the 
                     16-bp palindromic sequence 5'-CTGTATATATATACAG-3'."
                     /translation="MKALTARQQEVFDLIRDHISQTGMPPTRAEIAQRLGFRSPNAAEE
                     HLKALARKGVIEIVSGASRGIRLLQEEEEGLPLVGRVAAGEPLLAQQHIEGHYQVDPSL
                     FKPNADFLLRVSGMSMKDIGIMDGDLLAVHKTQDVRNGQVVVARIDDEVTVKRLKKQGN
                     KVELLPENSEFKPIVVDLRQQSFTIEGLAVGVIRNGDWL"
     protein_bind    6527..6596
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     primer_bind     complement(6636..6656)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(6636..6654)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6675..6691)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6675..6691)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6688..6710)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6699..6715
                     /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)."
     promoter        complement(join(6723..6729,6730..6747,6748..6753))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6768..6789
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
       61 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
      121 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
      181 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
      241 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
      301 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
      361 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      421 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      481 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
      541 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
      601 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      661 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
      721 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
      781 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
      841 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
      901 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
      961 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     1021 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     1081 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     1141 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     1201 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     1261 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1321 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     1381 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1441 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1501 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1561 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1621 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1681 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1741 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1801 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     1861 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     1921 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     1981 catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa aattcgcgtt
     2041 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     2101 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     2161 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     2221 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     2281 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     2341 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     2401 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     2461 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     2521 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     2581 gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat acgactcact
     2641 atagggcgaa ttggagctcg ctagcgatgt aggtcacggt ctcgaagccg cggtgcgggt
     2701 gccagggcgt gcccttgggc tccccgggcg cgtactccac ctcacccatc tggtccatca
     2761 tgatggtcga ctctagaagt cgatccaaca tggcgacttg tcccatcccc ggcatgttta
     2821 aatatactaa ttattcttga actaatttta atcaaccgat ttatctctct tccgcaggtc
     2881 gggaggttcc ggtggaagcg gaggtagcgg cggatccgag ggccgcggca gcctgctgac
     2941 ctgcggcgat gtggaggaga accccgggcc catgccaccc aagaagaagc gaaaagtaga
     3001 agatccaatg aaggctctca cggcccgaca acaggaagtt tttgatttga tacgggatca
     3061 tatatcccaa acgggtatgc ctccgacccg cgcagagata gcacagcgac tgggctttcg
     3121 atcgcctaac gccgcggagg agcacttgaa ggcactggcc cgcaagggtg tcattgaaat
     3181 cgtgtccggt gcgagccgcg gaatccggct gttgcaggag gaggaagagg gcctgccact
     3241 ggtgggacgc gtggccgctg gcgagccgct gctggcccag caacacatag agggacacta
     3301 tcaggtggac ccctccttgt ttaagccaaa tgctgatttc ctgttgcggg tgtcgggaat
     3361 gtccatgaag gacatcggta ttatggatgg tgacctcctc gccgtccata agacacagga
     3421 tgtcaggaac ggccaggtag tcgttgccag gatagacgat gaggtcactg tgaaacgtct
     3481 caagaagcaa ggcaataagg tcgagctgct gccggagaat agcgagttca agccgatcgt
     3541 ggtggatctg cgacagcagt cctttactat cgagggcttg gccgtgggtg tgatccgcaa
     3601 cggagattgg ctgggatcca tcgacagtgc ggcccaccac gacaacagca ccattccact
     3661 ggacttcatg cctcgtgacg cactgcatgg atttgactgg agcgaggagg atgatatgag
     3721 cgacggactc ccattcctga agaccgaccc caataacaat ggtttctttg gcgacggcag
     3781 cctgctgtgt atcctgcgaa gtattggttt taagccggaa aattacacga acagtaatgt
     3841 caatcgtctt cccacaatga tcaccgaccg ttacacgctg gccagtcgca gtactacaag
     3901 ccgcctgctg caaagctatc ttaacaactt ccacccctac tgccccatag tgcatagtcc
     3961 tactctcatg atgctttaca acaatcagat tgagatagca tcgaaggacc agtggcagat
     4021 cctcttcaac tgcattctcg ccatcggcgc ttggtgtatt gaaggcgaga gcacagatat
     4081 cgacgtgttt tactatcaaa acgccaagtc gcacctcacc tcgaaggtct tcgaaagtgg
     4141 aagcattatc ctcgtgacag ccctccatct gctgagtcgt tatacccagt ggcgtcagaa
     4201 aactaataca tcctacaact ttcattcgtt cagcattcgg atggcgatta gcctgggact
     4261 taatcgtgac ctcccaagta gcttcagcga ttcgtcgatc ctggaacagc gccggaggat
     4321 ttggtggagt gtgtacagct gggagatcca gctgtccctc ttgtacggac gcagcattca
     4381 gctgagtcag aacacgattt ccttcccgtc cagcgtggac gacgttcaac ggacaactac
     4441 gggcccgacc atctaccacg gtatcattga gacggcccgc ttgcttcaag tgtttacgaa
     4501 gatttatgag ctggataaga cagtgaccgc ggagaagagt ccgatctgcg ccaagaaatg
     4561 cctgatgatc tgcaatgaga tagaggaggt gagccgccag gcacccaagt tcctgcagat
     4621 ggacataagc actacggcgc tgactaatct cctcaaagaa cacccctggc tgagtttcac
     4681 ccgcttcgag ctgaagtgga aacagctgag tctgattata tacgttttgc gggacttttt
     4741 cacaaatttt acgcagaaga agtcgcagct tgagcaagat cagaacgacc atcagtccta
     4801 cgaggtaaag aggtgcagca taatgctctc cgacgcagcc cagcgcaccg tgatgtcggt
     4861 gtcgtcctat atggacaacc ataatgtgac cccttatttc gcctggaatt gctcctacta
     4921 tttgttcaac gccgtgttgg tcccaatcaa gactcttctg agcaactcca aatcgaacgc
     4981 cgagaataac gagacagcac agctcttgca gcaaattaac accgtgttga tgctgttgaa
     5041 gaaactcgct acattcaaga ttcagacttg cgaaaagtac atccaggtgt tggaagaggt
     5101 gtgcgcccca ttccttcttt cgcaatgcgc aattcccctc ccgcacataa gttacaataa
     5161 ttccaacggc tccgctatca agaatatcgt cggctccgct acaatcgcgc agtatcccac
     5221 gctgcctgag gaaaatgtta acaacatatc ggtgaagtat gtaagtccag gcagcgtcgg
     5281 cccgagccct gtgccgctga aaagcggagc ctccttcagc gatctggtca agctgctcag
     5341 caaccgccca ccatcccgca actcccctgt cactataccc cgatcgaccc cgtcgcatcg
     5401 tagcgttacc ccattcctgg gacaacagca gcaactgcag agtctggtgc cacttacgcc
     5461 ctcggcattg tttggcggag ccaacttcaa ccagagtggt aacattgcgg acagctcctt
     5521 gagcttcacg tttaccaaca gctccaacgg tccaaacttg atcacgactc agacaaactc
     5581 ccaggcattg tcccagccca ttgccagctc gaatgtgcat gataacttta tgaataatga
     5641 aattactgcc tccaagattg acgacggcaa caacagcaag cccctttccc caggctggac
     5701 tgaccagacg gcctacaatg ctttcggtat caccactggc atgttcaaca ccacaacgat
     5761 ggacgacgta tataactatc tttttgatga tgaggatact ccaccgaacc caaagaagga
     5821 gtagcctagg ttaattaata aaatgaatcg tagatactga aaaaccccgc aagttcactt
     5881 caactgtgca tcgtgcacca tctcaatttc tttcatttat acatcgtttt gccttctttt
     5941 atgtaactat actcctctaa gtttcaatct tggccatgta acctctgatc tatagaattt
     6001 tttaaatgac tagaattaat gcccatcttt tttttggacc taaattcttc atgaaaatat
     6061 attacgaggg cttattcaga agcttatcga taccgtcgac taaagccaaa tagaaattat
     6121 tcagttctgg cttaagtttt taaaagtgat attatttatt tggttgtaac caaccaaaag
     6181 aatgtaaata actaatacat aattatgtta gttttaagtt agcaacaaat tgattttagc
     6241 tatattagct acttggttaa taaatagaat atatttattt aaagataatt cgtttttatt
     6301 gtcagggagt gagtttgctt aaaaactcgt ttttaattaa agatccggag gtagcggtgg
     6361 aagcggaggt tccggcaggt aagttattga acaatggcat caaatgcctt catcatcact
     6421 accctttagc ccttaagacc ccacaatgac cttacccact cagagaaaaa agtaaatatg
     6481 aaagcccatt tgaacttcta agcttgtcga ccatcatgat ggaccagatg ggtgaggtgg
     6541 agtacgcgcc cggggagccc aagggcacgc cctggcaccc gcaccgcggc ttcgagaccg
     6601 tgacctacat catgcatggt acccagcttt tgttcccttt agtgagggtt aattgcgcgc
     6661 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     6721 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     6781 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgg
//
