LOCUS       sequence-416775-        6526 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG-tagged Drosophila IntS14 from inducible MtnA 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6526)
  AUTHORS   Fujiwara R, Zhai SN, Liang D, Shah AP, Tracey M, Ma XK, Fields CJ, 
            Mendoza-Figueroa MS, Meline MC, Tatomer DC, Yang L, Wilusz JE
  TITLE     IntS6 and the Integrator phosphatase module tune the efficiency of 
            select premature transcription termination events.
  JOURNAL   Mol Cell. 2023 Nov 14:S1097-2765(23)00902-4. doi: 
            10.1016/j.molcel.2023.10.035.
  PUBMED    37995689
REFERENCE   2  (bases 1 to 6526)
  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"; journalName: "Mol Cell. 
            2023 Nov 14:S1097-2765(23)00902-4. doi: 
            10.1016/j.molcel.2023.10.035."
COMMENT     Sequence Label: pMtnA FLAG-IntS14 puro
FEATURES             Location/Qualifiers
     source          1..6526
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     83..102
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     complement(716..732)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(716..732)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(729..751)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    740..756
                     /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(764..770,771..788,789..794))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    809..830
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(947..964)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1118..1706)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1198..1217)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1877..2668,2669..2737))
                     /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     2500..2519
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2738..2842)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2910..2928
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2966..2988)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3088..3107
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3301..3323
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3316..3332
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3338..3764
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     CDS             3789..3854
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             5663..5704
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             5714..5731
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5754..5771)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5773..5907
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5797..5816)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5851..5870
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(6029..6526,1..102))
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(6112..6132)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
ORIGIN
        1 gcgcgtggcg gggtagtcgg cgaacgcggc ggcgagggtg cgtacggccc gggggacgtc
       61 gtcgcgggtg gcgaggcgca ccgtgggctt gtactcggtc atggaaggtc gtctccttgt
      121 gaggggtcag gggcgtgggt caggggatgg tggcggcacc ggtcgtggcg gccgacctgc
      181 aggcatgcaa gctatccaat tcctgcagcc cgggggatct gttgtaattt ataatttata
      241 tttcctttct taataaataa ataaatagtc aagtttatgt ttgagtttta tgatttatat
      301 tttaagttat ttcaactgca acaccagcac cacgacctac ttacagcaaa aaacgtacaa
      361 gaaggaaaga aggaataaaa agagtggtat tctcttacaa tatgttttat ggcataaaag
      421 gtgtggccat tcatatcaaa tataaagtag tgttgtttaa cgttactttt gtaggttgaa
      481 tagtatattc caacagatga tgaggggttc ccaatcctaa acccatttgc cgttcccaga
      541 agcatgaaac caccacgcac ccgatcctct aggacaacaa caattgcatt cattttatgt
      601 ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg
      661 gtatggctga ttatgatcag tcgacctgca ggcatgcaag cttggcgtaa tcatggtcat
      721 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
      781 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
      841 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
      901 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
      961 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     1021 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     1081 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     1141 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     1201 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     1261 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     1321 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     1381 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     1441 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     1501 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     1561 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     1621 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     1681 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     1741 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     1801 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     1861 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     1921 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     1981 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     2041 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     2101 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     2161 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     2221 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     2281 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     2341 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     2401 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     2461 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     2521 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     2581 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     2641 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     2701 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     2761 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     2821 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     2881 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg
     2941 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     3001 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     3061 gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     3121 atatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgccatt
     3181 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     3241 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
     3301 cccagtcacg acgttgtaaa acgacggcca gtgaattaat tcgttgcagg acaggatgtg
     3361 gtgcccgatg tgactagctc tttgctgcag gccgtcctat cctctggttc cgataagaga
     3421 cccagaactc cggcccccca ccgcccaccg ccacccccat acatatgtgg tacgcaagta
     3481 agagtgcctg cgcatgcccc atgtgcccca ccaagagctt tgcatcccat acaagtcccc
     3541 aaagtggaga accgaaccaa ttcttcgcgg gcagaacaaa agcttctgca cacgtctcca
     3601 ctcgaatttg gagccggccg gcgtgtgcaa aagaggtgaa tcgaacgaaa gacccgtgtg
     3661 taaagccgcg tttccaaaat gtataaaacc gagagcatct ggccaatgtg catcagttgt
     3721 ggtcagcagc aaaatcaagt gaatcatctc agtgcaacta aaggggggat ctagatcggg
     3781 gtaccatgga ctacaaagac catgacggtg attataaaga tcatgatatc gattacaagg
     3841 atgacgatga caaggatcca ctagtcccca ccttaatagc gctggatgca tcgctatcga
     3901 tgctgcgccc ggtgccggga agaaatgagc acacctacca gtcgctggcc accaagggca
     3961 tccagcatct gctggacaat ctcacggcgg ccggcaaatt ggagcacgtt gctctgctct
     4021 cctattcgac gacggcggaa ctgaaggtgg acttcacgcg ggactacgac caggtgcggc
     4081 aggcggtcaa gaaggtggag cccgtggaca aggcctgcct gatgagcatg ctcaaggcgg
     4141 tggtgtccat aatgtcgccg tggggcaacc agaacatcct gcaggtggtt gtctttacgg
     4201 actgcggcct cggctttggc aacacctcga tcacgggctt cctggaggcc tacgccgaaa
     4261 aggagtcgga gccggagttc ggcttcctga agactctggc caactacaac ctgaatttca
     4321 tctgcctggg cctgcacggc gattactact tcaccagggg attggcggtg taccagcagc
     4381 tgctggacaa ggtctcactg aagggccagc tcttcatgac aaagcccgcc aagagcagcg
     4441 atgcagtcga gggaaatccc aatcccaacc cgaaccccag ccacaaaagc gaactgggcc
     4501 gcactacggt cttcgaacta attgaacgtc tctgcgaggc cagctacaag agttcggagg
     4561 tgaccctgaa atgcggcagc tacttccgca tggaggcatc tgttctgctc tggccaccga
     4621 cggctccgta tgagcaaaag tcccacatat tcggccgaga acccaccatt cgccacacag
     4681 accagaagat tgaggtgtgt ggcttcctgt ccctttcgga cattggatca ccggccaccc
     4741 tgagtcgaca ctgggtgctg cccaaagtgg agcgggagaa gagcggcagc agtcggaggt
     4801 ctggaaacct gagcgcagca gccaagccac cgaagctcaa tctagatacc agcaatccca
     4861 actacgagct ggagaagctg gaagccgata tcaaagagtt ttacgccaag gattcaaaag
     4921 acacggagga aagtggcgac gacgatgtga ccattgtctt gaaacccggc ccccagacag
     4981 agcaacaaaa agaaaacttg tgcgtcctcc ttcacggagc cctcaaaatg gagaacatgg
     5041 cagctctggt tcgagtgggt gacaagtggt acggattcat atatgcattc accgacagca
     5101 agaagaagtc aaacctgatg ctgaacatac ttccacccgg cacgaacgtg attccctggc
     5161 taggagacct ggaatcgctg ggcttccccg aggatttggc gcccggagag acggccagtt
     5221 ttccggtgcg tgccgaccgc agatcttatt cgcagagcag tgtggtgtgg atacgacagg
     5281 ctagcctgca gtccgacgtc cagaaggtac tgcgacacgc caagaagatg cccgacaaga
     5341 cgcagcactt ctacaaggag ctcaatcgca tccgccgcgc tgccctggcg ctgggattcg
     5401 tcgagttgct ggaggctttg gccatgctgc tggagaagga gtgcgcccat ctcagcctga
     5461 acggagccag caacgactgc accctgcagc tgcagcacgc ggctacggag ttgcggaaga
     5521 cctcgaacag agacatgaag tccatgattg ttccgctcca gaaagttggt gcatcggatg
     5581 cgggcagcac ggctgccacg gctcctgctc ctgcatacat gtattgagcg gccgctcgag
     5641 tctagagggc ccgcggttcg aaggtaagcc tatccctaac cctctcctcg gtctcgattc
     5701 tacgcgtacc ggtcatcatc accatcacca ttgagtttaa acccgctgat cagcctcgac
     5761 tgtgccttct aagatccaga catgataaga tacattgatg agtttggaca aaccacaact
     5821 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
     5881 accattataa gctgcaataa acaagttcta gggtcggtgg gcctcggggg cgggtgcggg
     5941 gtcggcgggg ccgccccggg tggcttcggt cggagccatg gggtcgtgcg ctcctttcgg
     6001 tcgggcgctg cgggtcgtgg ggcgggcgtc aggcaccggg cttgcgggtc atgcaccagg
     6061 tgcgcggtcc ttcgggcacc tcgacgtcgg cggtgacggt gaagccgagc cgctcgtaga
     6121 aggggaggtt gcggggcgcg gaggtctcca ggaaggcggg caccccggcg cgctcggccg
     6181 cctccactcc ggggagcacg acggcgctgc ccagaccctt gccctggtgg tcgggcgaga
     6241 cgccgacggt ggccaggaac cacgcgggct ccttgggccg gtgcggcgcc aggaggcctt
     6301 ccatctgttg ctgcgcggcc agccgggaac cgctcaactc ggccatgcgc gggccgatct
     6361 cggcgaacac cgcccccgct tcgacgctct ccggcgtggt ccagaccgcc accgcggcgc
     6421 cgtcgtccgc gacccacacc ttgccgatgt cgagcccgac gcgcgtgagg aagagttctt
     6481 gcagctcggt gacccgctcg atgtggcggt ccgggtcgac ggtgtg
//
