LOCUS       sequence-104087-        8748 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Creating Gal80 lines from MiMIC lines with inserts into coding 
            introns in Phase 0. Gal80 is tagged with FLAG, HA and EGFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8748)
  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 8748)
  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(0)-T2A-3X Gal80-Hsp70
FEATURES             Location/Qualifiers
     source          1..8748
                     /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             2917..2970
                     /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             4276..4302
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4306..4329
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4330..4386
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5692..6405
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             6406..6459
                     /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"
     protein_bind    8452..8521
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     primer_bind     complement(8561..8581)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(8561..8579)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(8600..8616)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8600..8616)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8613..8635)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8624..8640
                     /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(8648..8654,8655..8672,8673..8678))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8693..8714
                     /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 tccgcaggtg
     2881 ggaggttccg gtggaagcgg aggtagcggc ggatccgagg gccgcggcag cctgctgacc
     2941 tgcggcgatg tggaggagaa ccccgggccc atggattaca acaaaaggag tagtgtgagt
     3001 acggtgccga atgctgctcc cattcgcgtg ggcttcgtgg gattgaacgc ggctaagggt
     3061 tgggccatta aaacgcatta tccagccata ctgcagctga gctcccagtt ccaaataaca
     3121 gcgctttatt cccccaagat cgagacgtcc attgcgacaa ttcagaggct gaagttgtcc
     3181 aacgccacag ctttcccaac actcgagagc ttcgcctcga gctccacgat cgatatgatt
     3241 gtgatcgcaa tccaggtggc ttcccactac gaggtggtaa tgcccctgct cgagttttcg
     3301 aagaacaatc ctaatctgaa gtatttgttt gtggaatggg cactggcctg ctcgctcgat
     3361 caggcggagt cgatctataa ggccgctgca gagcggggcg tgcaaacaat aatcagtctg
     3421 cagggacgaa agagcccgta cattttgcgg gcaaaagagc ttatctccca gggctacatc
     3481 ggcgatataa atagcatcga gattgcaggt aacggaggtt ggtacggtta cgagcggccg
     3541 gtgaaaagcc cgaaatacat ttacgagatc ggaaacggag ttgatctggt gaccaccacg
     3601 ttcggtcaca cgatagatat attgcagtac atgaccagtt cgtacttcag ccgtatcaat
     3661 gccatggtgt ttaacaacat tccagaacag gagctcattg atgagagggg caatcgcctg
     3721 ggccagcggg tcccaaagac tgtcccagat catctcttgt tccagggcac gctgctcaac
     3781 ggaaatgttc ctgtgtcctg ttcctttaag ggaggcaagc ctacgaaaaa gttcaccaag
     3841 aatctggtca tagatatcca tggtacaaag ggtgatctga agctggaggg tgacgctgga
     3901 tttgcggaaa tctcgaatct cgttctgtat tattcgggca cccgcgccaa tgattttccc
     3961 ctggccaatg gtcagcaggc acccttggac cccggctacg acgcaggcaa ggagattatg
     4021 gaggtgtatc acctccgaaa ctacaacgcg atcgttggta atatacaccg gctctaccag
     4081 agtatcagcg atttccactt caacaccaag aaaatccccg agcttccatc gcagttcgta
     4141 atgcaaggct tcgatttcga aggctttccc accctgatgg atgccttgat cttgcatcgc
     4201 ctgattgaaa gtgtctacaa gtcgaatatg atgggcagta ccctgaatgt ttccaacatc
     4261 tcccactact cgctttaccc ctacgatgtg cccgattacg ccggcgatta caaggacgat
     4321 gacgacaagg ccaccaactt cagcctgctg aagcaggccg gcgatgtgga ggagaacccc
     4381 gggcccatgg attacaacaa aaggagtagt gtgagtacgg tgccgaatgc tgctcccatt
     4441 cgcgtgggct tcgtgggatt gaacgcggct aagggttggg ccattaaaac gcattatcca
     4501 gccatactgc agctgagctc ccagttccaa ataacagcgc tttattcccc caagatcgag
     4561 acgtccattg cgacaattca gaggctgaag ttgtccaacg ccacagcttt cccaacactc
     4621 gagagcttcg cctcgagctc cacgatcgat atgattgtga tcgcaatcca ggtggcttcc
     4681 cactacgagg tggtaatgcc cctgctcgag ttttcgaaga acaatcctaa tctgaagtat
     4741 ttgtttgtgg aatgggcact ggcctgctcg ctcgatcagg cggagtcgat ctataaggcc
     4801 gctgcagagc ggggcgtgca aacaataatc agtctgcagg gacgaaagag cccgtacatt
     4861 ttgcgggcaa aagagcttat ctcccagggc tacatcggcg atataaatag catcgagatt
     4921 gcaggtaacg gaggttggta cggttacgag cggccggtga aaagcccgaa atacatttac
     4981 gagatcggaa acggagttga tctggtgacc accacgttcg gtcacacgat agatatattg
     5041 cagtacatga ccagttcgta cttcagccgt atcaatgcca tggtgtttaa caacattcca
     5101 gaacaggagc tcattgatga gaggggcaat cgcctgggcc agcgggtccc aaagactgtc
     5161 ccagatcatc tcttgttcca gggcacgctg ctcaacggaa atgttcctgt gtcctgttcc
     5221 tttaagggag gcaagcctac gaaaaagttc accaagaatc tggtcataga tatccatggt
     5281 acaaagggtg atctgaagct ggagggtgac gctggatttg cggaaatctc gaatctcgtt
     5341 ctgtattatt cgggcacccg cgccaatgat tttcccctgg ccaatggtca gcaggcaccc
     5401 ttggaccccg gctacgacgc aggcaaggag attatggagg tgtatcacct ccgaaactac
     5461 aacgcgatcg ttggtaatat acaccggctc taccagagta tcagcgattt ccacttcaac
     5521 accaagaaaa tccccgagct tccatcgcag ttcgtaatgc aaggcttcga tttcgaaggc
     5581 tttcccaccc tgatggatgc cttgatcttg catcgcctga ttgaaagtgt ctacaagtcg
     5641 aatatgatgg gcagtaccct gaatgtttcc aacatctccc actactcgct tatgtccaaa
     5701 ggtgaagaac tgtttaccgg agtagtcccg atattggttg aactcgacgg cgatgtcaac
     5761 ggtcataaat tcagtgtgtc cggcgagggt gagggcgacg ccacatacgg taagctgacg
     5821 ttgaagttca tatgcaccac gggcaagctg cccgtgccat ggccgacgtt ggtcacgacg
     5881 ctgacgtatg gtgtccagtg tttcagccgt taccccgatc atatgaagca gcatgacttt
     5941 ttcaagtcgg cgatgccgga gggatacgtt caagagagga ccattttctt caaggatgac
     6001 ggcaactata agacgcgagc ggaggtgaaa tttgaaggcg acacactcgt taaccgtatt
     6061 gagttgaagg gcattgattt taaggaggat ggcaacattc tgggccataa gttggaatac
     6121 aactacaact cgcataatgt gtatataatg gcagataagc agaaaaacgg aataaaggtt
     6181 aacttcaaga ttcgccacaa catagaggac ggttccgtgc aacttgcaga tcattaccaa
     6241 cagaacacac ccattggaga tggcccagtt ctcttgccag acaatcacta cctttccaca
     6301 cagtccgcgt tgagcaagga ccccaatgaa aagcgggacc acatggtgtt gctggagttt
     6361 gtgaccgcag ctggtattac acacggcatg gatgagctct acaaggaggg ccgcggcagc
     6421 ctgctgacct gcggcgatgt ggaggagaac cccgggccca tggattacaa caaaaggagt
     6481 agtgtgagta cggtgccgaa tgctgctccc attcgcgtgg gcttcgtggg attgaacgcg
     6541 gctaagggtt gggccattaa aacgcattat ccagccatac tgcagctgag ctcccagttc
     6601 caaataacag cgctttattc ccccaagatc gagacgtcca ttgcgacaat tcagaggctg
     6661 aagttgtcca acgccacagc tttcccaaca ctcgagagct tcgcctcgag ctccacgatc
     6721 gatatgattg tgatcgcaat ccaggtggct tcccactacg aggtggtaat gcccctgctc
     6781 gagttttcga agaacaatcc taatctgaag tatttgtttg tggaatgggc actggcctgc
     6841 tcgctcgatc aggcggagtc gatctataag gccgctgcag agcggggcgt gcaaacaata
     6901 atcagtctgc agggacgaaa gagcccgtac attttgcggg caaaagagct tatctcccag
     6961 ggctacatcg gcgatataaa tagcatcgag attgcaggta acggaggttg gtacggttac
     7021 gagcggccgg tgaaaagccc gaaatacatt tacgagatcg gaaacggagt tgatctggtg
     7081 accaccacgt tcggtcacac gatagatata ttgcagtaca tgaccagttc gtacttcagc
     7141 cgtatcaatg ccatggtgtt taacaacatt ccagaacagg agctcattga tgagaggggc
     7201 aatcgcctgg gccagcgggt cccaaagact gtcccagatc atctcttgtt ccagggcacg
     7261 ctgctcaacg gaaatgttcc tgtgtcctgt tcctttaagg gaggcaagcc tacgaaaaag
     7321 ttcaccaaga atctggtcat agatatccat ggtacaaagg gtgatctgaa gctggagggt
     7381 gacgctggat ttgcggaaat ctcgaatctc gttctgtatt attcgggcac ccgcgccaat
     7441 gattttcccc tggccaatgg tcagcaggca cccttggacc ccggctacga cgcaggcaag
     7501 gagattatgg aggtgtatca cctccgaaac tacaacgcga tcgttggtaa tatacaccgg
     7561 ctctaccaga gtatcagcga tttccacttc aacaccaaga aaatccccga gcttccatcg
     7621 cagttcgtaa tgcaaggctt cgatttcgaa ggctttccca ccctgatgga tgccttgatc
     7681 ttgcatcgcc tgattgaaag tgtctacaag tcgaatatga tgggcagtac cctgaatgtt
     7741 tccaacatct cccactactc gctttaataa taaaatgaat cgtagatact gaaaaacccc
     7801 gcaagttcac ttcaactgtg catcgtgcac catctcaatt tctttcattt atacatcgtt
     7861 ttgccttctt ttatgtaact atactcctct aagtttcaat cttggccatg taacctctga
     7921 tctatagaat tttttaaatg actagaatta atgcccatct tttttttgga cctaaattct
     7981 tcatgaaaat atattacgag ggcttattca gaagcttatc gataccgtcg actaaagcca
     8041 aatagaaatt attcagttct ggcttaagtt tttaaaagtg atattattta tttggttgta
     8101 accaaccaaa agaatgtaaa taactaatac ataattatgt tagttttaag ttagcaacaa
     8161 attgatttta gctatattag ctacttggtt aataaataga atatatttat ttaaagataa
     8221 ttcgttttta ttgtcaggga gtgagtttgc ttaaaaactc gtttggatcc ggaggtagcg
     8281 gtggaagcgg aggttccggc gaggtaagtt attgaacaat ggcatcaaat gccttcatca
     8341 tcactaccct ttagccctta agaccccaca atgaccttac ccactcagag aaaaaagtaa
     8401 atatgaaagc ccatttgaac ttctaagctt gtcgaccatc atgatggacc agatgggtga
     8461 ggtggagtac gcgcccgggg agcccaaggg cacgccctgg cacccgcacc gcggcttcga
     8521 gaccgtgacc tacatcatgc atggtaccca gcttttgttc cctttagtga gggttaattg
     8581 cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     8641 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     8701 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcgg
//
