LOCUS       sequence-383956-        8610 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses FLAG-tagged Drosophila IntS6 from inducible MtnA promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8610)
  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 8610)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-IntS6 puro
FEATURES             Location/Qualifiers
     source          1..8610
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(97..116)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      910..1498
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1399..1418
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1652..1669
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1786..1807
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1822..1827,1828..1845,1846..1852)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1860..1876
                     /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)."
     primer_bind     1865..1887
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1884..1900
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1884..1900
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             2514..3113
                     /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(2514..2533)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3010..3030
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3235..3369
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3272..3291)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3326..3345
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3371..3388
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(3411..3428)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(3438..3479)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(7372..7437)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     promoter        complement(7462..7888)
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     primer_bind     complement(7894..7911)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7894..7910)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7903..7925)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(8119..8138)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8238..8260
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8298..8316)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8384..8488
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8489..8557,8558..8610,1..739)
                     /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"
ORIGIN
        1 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
       61 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
      121 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
      181 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
      241 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
      301 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
      361 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
      421 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
      481 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
      541 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
      601 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
      661 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
      721 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
      781 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
      841 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
      901 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
      961 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     1021 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     1081 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     1141 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     1201 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     1261 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     1321 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     1381 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     1441 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     1501 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     1561 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     1621 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     1681 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
     1741 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
     1801 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
     1861 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
     1921 atgcctgcag gtcgactgat cataatcagc cataccacat ttgtagaggt tttacttgct
     1981 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
     2041 gtcctagagg atcgggtgcg tggtggtttc atgcttctgg gaacggcaaa tgggtttagg
     2101 attgggaacc cctcatcatc tgttggaata tactattcaa cctacaaaag taacgttaaa
     2161 caacactact ttatatttga tatgaatggc cacacctttt atgccataaa acatattgta
     2221 agagaatacc actcttttta ttccttcttt ccttcttgta cgttttttgc tgtaagtagg
     2281 tcgtggtgct ggtgttgcag ttgaaataac ttaaaatata aatcataaaa ctcaaacata
     2341 aacttgacta tttatttatt tattaagaaa ggaaatataa attataaatt acaacagatc
     2401 ccccgggctg caggaattgg atagcttgca tgcctgcagg tcggccgcca cgaccggtgc
     2461 cgccaccatc ccctgaccca cgcccctgac ccctcacaag gagacgacct tccatgaccg
     2521 agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt cccccgggcc gtacgcaccc
     2581 tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgacccg gaccgccaca
     2641 tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca
     2701 aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg
     2761 aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc
     2821 tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt
     2881 ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg
     2941 tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct
     3001 ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg
     3061 aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgacgcccgc
     3121 cccacgaccc gcagcgcccg accgaaagga gcgcacgacc ccatggctcc gaccgaagcc
     3181 acccggggcg gccccgccga ccccgcaccc gcccccgagg cccaccgacc ctagaacttg
     3241 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     3301 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     3361 gtctggatct tagaaggcac agtcgaggct gatcagcggg tttaaactca atggtgatgg
     3421 tgatgatgac cggtacgcgt agaatcgaga ccgaggagag ggttagggat aggcttacct
     3481 tcgaaccgcg ggccctctag actcgagtta actcttggcg acggcctgct ccgcgctggg
     3541 ggtcgcttca ttggttccca cggcagttaa ctgctgcttg cgatcccacc actcctgtat
     3601 gctggacgcc atgcgatggc ggcggaagcg cagcgattcc acgatacaca tgtccacgaa
     3661 atccgactgc atatcctggt cgcccttgac cagattcagg tgctccagca gctggctgta
     3721 gtctcggccg ggtcgccgta tatcacggaa gatctgcaat cgtagttcga cgttgtgcaa
     3781 ccgggccgct tcgcgctgct cctcggtgaa aatgagcggc aggctctgtc ccggtgagcc
     3841 catgcccccg ttgttgtgcg tcggtgagga gccgcagctg gagttgatct tcacctccag
     3901 tttgttgggg ctgctgttgt tgtgaccccg gtggttgtga ttgttgctca acgtcgaacc
     3961 gctagctgct ccagcggcgc tgctgttgtt attgttgtga tgattgagac cactggccca
     4021 gtagtagtgg ggatttcccg ctgcagtgct gcccgcagta cccgctgtcc gtttcagggc
     4081 atcgaaactg ttgccaccca gcgaggcatg atcgtcggat gagactcctc cgccactctc
     4141 cgtgctgcca ttgccattca aattgctgtt gcttgcgccc gctccgaccg aggagccact
     4201 gctggttcca ttgtgacagg tctgcagtat gcgcgagatc tccgaggcat cgttgatgtc
     4261 attgtggctg aagtaggcgg aactagacga tagtcccgag ctgctggcag cgggcgtact
     4321 tgttcccaat ggtgatagag gcaccaagga caatggcgaa ggtggggaaa tggctgaact
     4381 atgaccgtgt ccataaccat tggccagtgg cagcgatccc ggcttgctga caccgagtag
     4441 tcccgatcct ccgccacccg ccgctccacc cagggagcta aggttgtcca acggcgagct
     4501 cggaccaccc agattactgc tgtaggccgc atgggtggtc agtccgttga ggggcacgta
     4561 gagaacgccc ggggaactgg cgctggagct gatggaactg ctgctgccgg ccgcgacggc
     4621 ggagcaaact tcgctgccat tgctggcggg gatcacagat gttgctggcg tggctgaaac
     4681 tgctggtgat gctccaattg acgacggttc gttagctgaa gacgcacccg gcagggatgc
     4741 cccgcctgga actgcttccg tttcgcccgt ttctgcatcg tcgctgatgc cattgatgaa
     4801 attgtgcaca taaccattga gcggcgtatc ctgatcgctg gtctcctcct cgatgaaaac
     4861 gaaactgatt cgcaagggat ccaaatgcga gtttgacatc cctgaatcac tggccgaggc
     4921 ctccgatgtg gagctcacga agctgctgtt gctattgctg tcctcgtcga aacagctgga
     4981 actgggaccg ccatcagcgc ccatggcgga atcgcagccg gatatggatg acgtggcgcc
     5041 tggtatcggt gatcctggtg cactacccgt tgtggataat tcggagccgg agctcgaaga
     5101 tcgcgagtcc gtagatgatc gacggaattc aaagctacgt ggcagcgggc ctgctttcct
     5161 ctttcgccca cgagacgtgc cgtccagttt tttagaggat ccgccactgc cacctgcgcc
     5221 cagcatcagt cctgacattc caccgcccat tccgggcatt ccgggcatcc cgggacccga
     5281 accaccgccg ggcgaagagc ccggagatga gggacccggt cctgtgggcg agcccgatcc
     5341 ggaaggtgag ccgggtggcg ctccacccgg tccattgcca tttggcgact tcattggcgc
     5401 cacatcgctc agatccgcct cgtccaccat caccatgtgc ttgttcttct tgtacggatt
     5461 accgaacatg tgctgccgca cattcgtcgg ctcgatttca cgcagcggat tgtccttgtt
     5521 cttgaggtac tcctggtagt tgcccatctc ggcgatgggc aggcaatggc ccgagtcctt
     5581 ggccaccagc gagacgggtc gcagtaaggt ctcacgcatc ttggctattt cctccaccaa
     5641 atcccgccgc ggtatatcga agggattgcg atgcgacttg gccgacgact gatgcgttgg
     5701 ctgcggcacc acaatcgtat aattctccac gggatctatt tccccatgca acttggaaaa
     5761 tatatcccgg agcagtggat gccgcaccaa atctctgcgt aacggcgcac ctgtgaacag
     5821 tttgcccgtc tccacttggc gatatggtgg cttcggctgc ttcagctgct tgtacacctt
     5881 catgcaaaga ttctcctgat cctgcttggc cgtattcttg atgtgcttaa gctgattggc
     5941 gatcgttggc gacagatagt tgtccacatt ctccggcagc agaaactgca gcagctgata
     6001 cggcacattg atgttgacca gcgccttgcg caggaacggg cagtaatacg ggggtattga
     6061 acgcacatag gcgttgaatt tgtacatcag atcgctcggc ggactcatat tgtacttgtg
     6121 aatgaggtca tgcagaatgg gcagcaacgc tggataattg taggccagta catagaggtg
     6181 tacctgcgaa aagttgggcg ccgcttttag gtagccaaag ggcagctcga atccatgcat
     6241 gccattggtc acaatcacct gccagcactt gttcatctcg cgcttgttga gaatctgcag
     6301 cgttaacgga cagccttcaa tctcgtactt gtcgacgggg aagctgcgca ccagctgtgg
     6361 ctcgtccacc gccggcgtca ggaccttcag cttgggatga gcgtcgcgag gcggcaatgt
     6421 aatcgccttg gagtccggcc aatagggctc cggcagtggc caatagccga tgggaaaggt
     6481 tttctgcgtg atgtgcttct gcacgtagat catcttcttt accggatgga agacaatatc
     6541 gggagcggga cccgctccgg acgttgaggg tattcccatg ccggatatcg tgctcgcacc
     6601 cgccgcttcg ccagcggtag cggaagtagc ctccttgggc agcataggct cgaactgcag
     6661 caccacacca ggctgaacct tttggacaag actttcaatg cactggttga gtacgtagtg
     6721 gcttcgcact cgatatgaac gtccgccggt aacctcgcac atccgttcga tgggagaatc
     6781 gtcatgcggc accttgccat ccactcgctc gtcaatcttg ttgcccggca tgcggaggac
     6841 aagcgaaaac aagcgctgat cccagcgaaa tggctcctta gtgaactttg tgcccggtat
     6901 ttggttactt agcggcagta tgatctcctg atggacacca ttccggtacg aatagcgacc
     6961 gccgtccgta atcacaatga tgaccgatgg ctccagataa aacgggcacc tgccctgccc
     7021 gtacgtatcg atgcccgact gcatgcgatt cagatttaac aaatcgaacg cattgcgcag
     7081 cgattcaccc atcgaggtga ggccgtgact ctgcaggttc ttcagctcgt tcatgaaggt
     7141 ggcatggttc tccttccatc cagctttcac gtttgccggt ggctcctcga atgtgagcag
     7201 catgtagcga tctcccaggc aatcctgcgt acgctggcga tacttgagaa acgtctccac
     7261 ggctcccttg gcaatgtcca gatacgtttt ctgtacccca ttcacatacg ccttctggca
     7321 catggacgac gaggtgtcca ccaggaagag tatgattgtg actagtggat ccttgtcatc
     7381 gtcatccttg taatcgatat catgatcttt ataatcaccg tcatggtctt tgtagtccat
     7441 ggtaccccga tctagatccc ccctttagtt gcactgagat gattcacttg attttgctgc
     7501 tgaccacaac tgatgcacat tggccagatg ctctcggttt tatacatttt ggaaacgcgg
     7561 ctttacacac gggtctttcg ttcgattcac ctcttttgca cacgccggcc ggctccaaat
     7621 tcgagtggag acgtgtgcag aagcttttgt tctgcccgcg aagaattggt tcggttctcc
     7681 actttgggga cttgtatggg atgcaaagct cttggtgggg cacatggggc atgcgcaggc
     7741 actcttactt gcgtaccaca tatgtatggg ggtggcggtg ggcggtgggg ggccggagtt
     7801 ctgggtctct tatcggaacc agaggatagg acggcctgca gcaaagagct agtcacatcg
     7861 ggcaccacat cctgtcctgc aacgaattaa ttcactggcc gtcgttttac aacgtcgtga
     7921 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     7981 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     8041 tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     8101 catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     8161 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     8221 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     8281 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     8341 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     8401 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     8461 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     8521 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     8581 aaaagatgct gaagatcagt tgggtgcacg
//
