LOCUS       sequence-258481-        7755 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses nanoLuciferase (nLuc) mRNA from the Drosophila 
            Metallothionein A promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7755)
  AUTHORS   Tatomer DC, Elrod ND, Liang D, Xiao MS, Jiang JZ, Jonathan M, Huang 
            KL, Wagner EJ, Cherry S, Wilusz JE
  TITLE     The Integrator complex cleaves nascent mRNAs to attenuate 
            transcription.
  JOURNAL   Genes Dev. 2019 Sep 17. pii: gad.330167.119. doi: 
            10.1101/gad.330167.119.
  PUBMED    31530651
REFERENCE   2  (bases 1 to 7755)
  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: "Genes Dev. 
            2019 Sep 17. pii: gad.330167.119. doi: 10.1101/gad.330167.119."
COMMENT     Sequence Label: Hy_pMtnA nLuc SV40
FEATURES             Location/Qualifiers
     source          1..7755
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          307..372
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             502..522
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    947..1081
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1082..1508
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     primer_bind     1516..1532
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1517..1533
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1551..2063
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             2064..2129
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    2141..2262
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2972..2989)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3143..3731)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3223..3242)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3902..4693,4694..4762))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4525..4544
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4763..4867)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4935..4953
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6349..6628
                     /label=copia promoter
                     /note="strong promoter from the Drosophila transposable 
                     element copia (Sinclair et al., 1986)"
     CDS             6674..7699
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMPRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
ORIGIN
        1 gcgagcctag caaggcaaat gcgagagaac ggccttacgc ttggtggcac agttctcgtc
       61 cacagttcgc taagctcgct cggctgggtc gcgggagggc cggtcgcagt gattcaggcc
      121 cttctggatt gtgttggtcc ccagggcacg attgtcatgc ccacgcactc gggtgatctg
      181 actgatcccg cagattggag atcgccgccc gtgcctgccg attgggtgca gatctttgtg
      241 aaggaacctt acttctgtgg tgtgacataa ttggacaaac tacctacaga gatttaaagc
      301 tctaaggtaa atataaaatt tttaagtgta taatgtgtta aactactgat tctaattgtt
      361 tgtgtatttt agattccaac ctatggaact gatgaatggg agcagtggtg gaatgccttt
      421 aatgaggaaa acctgttttg ctcagaagaa atgccatcta gtgatgatga ggctactgct
      481 gactctcaac attctactcc tccaaaaaag aagagaaagg tagaagaccc caaggacttt
      541 ccttcagaat tgctaagttt tttgagtcat gctgtgttta gtaatagaac tcttgcttgc
      601 tttgctattt acaccacaaa ggaaaaagct gcactgctat acaagaaaat tatggaaaaa
      661 tattctgtaa cctttataag taggcataac agttataatc ataacatact gttttttctt
      721 actccacaca ggcatagagt gtctgctatt aataactatg ctcaaaaatt gtgtaccttt
      781 agctttttaa tttgtaaagg ggttaataag gaatatttga tgtatagtgc cttgactaga
      841 gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca
      901 cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc
      961 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     1021 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggat
     1081 caattcgttg caggacagga tgtggtgccc gatgtgacta gctctttgct gcaggccgtc
     1141 ctatcctctg gttccgataa gagacccaga actccggccc cccaccgccc accgccaccc
     1201 ccatacatat gtggtacgca agtaagagtg cctgcgcatg ccccatgtgc cccaccaaga
     1261 gctttgcatc ccatacaagt ccccaaagtg gagaaccgaa ccaattcttc gcgggcagaa
     1321 caaaagcttc tgcacacgtc tccactcgaa tttggagccg gccggcgtgt gcaaaagagg
     1381 tgaatcgaac gaaagacccg tgtgtaaagc cgcgtttcca aaatgtataa aaccgagagc
     1441 atctggccaa tgtgcatcag ttgtggtcag cagcaaaatc aagtgaatca tctcagtgca
     1501 actaaagggg ggatctcgag gtcgacggta tcgataagct tgatatcacc atggtcttca
     1561 cactcgaaga tttcgttggg gactggcgac agacagccgg ctacaacctg gaccaagtcc
     1621 ttgaacaggg aggtgtgtcc agtttgtttc agaatctcgg ggtgtccgta actccgatcc
     1681 aaaggattgt cctgagcggt gaaaatgggc tgaagatcga catccatgtc atcatcccgt
     1741 atgaaggtct gagcggcgac caaatgggcc agatcgaaaa aatttttaag gtggtgtacc
     1801 ctgtggatga tcatcacttt aaggtgatcc tgcactatgg cacactggta atcgacgggg
     1861 ttacgccgaa catgatcgac tatttcggac ggccgtatga aggcatcgcc gtgttcgacg
     1921 gcaaaaagat cactgtaaca gggaccctgt ggaacggcaa caaaattatc gacgagcgcc
     1981 tgatcaaccc cgacggctcc ctgctgttcc gagtaaccat caacggagtg accggctggc
     2041 ggctgtgcga acgcattctg gcggactaca aagaccatga cggtgattat aaagatcatg
     2101 acatcgatta caaggatgac gatgacaagt aagcggccgc aacttgttta ttgcagctta
     2161 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     2221 gcattctagt tgtggtttgt ccaaactcat caatgtatct taactagtgg atccactagg
     2281 ggccgccgac gcgaggctgg atggccttcc ccattatgat tcttctcgct tccggcggca
     2341 tcgggatgcc cgcgttgcag gccatgctgt ccaggcaggt agatgacgac catcagggac
     2401 agcttcaagg atcgctcgcg gctcttacca gcctaacttc gatcattgga ccgctgatcg
     2461 tcacggcgat ttatgccgcc tcggcgagca catggaacgg gttggcatgg attgtaggcg
     2521 ccgccctata ccttgtctgc ctccccgcgt tgcgtcgcgg tgcatggagc cgggccacct
     2581 cgacctgaat ggaagccggc ggcacctcgc taacggattc accactccaa gaattggagc
     2641 caatcaattc ttgcggagaa ctgtgaatgc gcaaaccaac ccttggcaga acatatccat
     2701 cgcgtccgcc atctccagca gccgcacgcg gcgcatctcg ggcagcgttg ggtcctggcc
     2761 acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc ggggttgcct
     2821 tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact gctgctgcaa
     2881 aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt cgtaaagtct
     2941 ggaaacgcgg aagtcagcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     3001 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     3061 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     3121 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     3181 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     3241 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     3301 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     3361 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     3421 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3481 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     3541 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     3601 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     3661 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     3721 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     3781 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     3841 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     3901 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     3961 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     4021 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     4081 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     4141 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     4201 acgttgttgc cattgctgca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     4261 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     4321 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     4381 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     4441 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     4501 gctcttgccc ggcgtcaaca cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     4561 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     4621 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     4681 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     4741 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     4801 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     4861 ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga
     4921 cattaaccta taaaaatagg cgtatcacga ggccctttcg tcttcaagaa ttcacatttg
     4981 tacgaatttt ttttttatca aaagttcgag tttttcacca atttcctcat caaccgagca
     5041 aggcaaacgg ctttgaataa tatggtgtta tatatacata tatcaaatcg ctgctgactg
     5101 cgtgattgat ggccccaaga ttacatatta tcgaatcagg attcagaagg agatcaatgt
     5161 caaatgcgga caggaacatg aaagacgcct gttatgcgca attaaaaatt tgggtttaat
     5221 tgctgtggaa actgttgttg gcggcatctt aagttcctgt ttaacaacat caactactta
     5281 tgtacgtaga agcgtttaag ccatttgcat acagatgaga actggctttt gtgctaatca
     5341 gtcaagatga ctccgatgat gatgactcat tacctgacca gttttcgctg ctttcttttc
     5401 aacaactact tgtatatgta ttgtatccaa tagcaataca ttgaatttcc atggtctagt
     5461 cacgtattat catttaattg acaccaagtc gtgttattgt tgagctatcg agttcagctc
     5521 aaacatttct tattcccatg aataagccgg caaaaatatg caatctatga aagttaatat
     5581 aagcaaacct tactttgact caataccaat gcactttgtg tcgataggtt cacgcaattg
     5641 aggcgattat tccgataacc caagcgattg actgttcccg tttcgattcc aattgaaatt
     5701 tggaaatgta caatagtttt gctatatgct gtcaagtacg ctcttatctt ctctgggttt
     5761 tcttcagagt ttcgaaacgc ttcttctttt ttttgttttt ttttttttgg aatctcgtat
     5821 tttggaaggg gctcccctct ggaatttgtt acactgtcgt tatcattgcg aacaagcggc
     5881 ccgaagctat cagcgacttt aacatttaca atgcactttt ttacgaccaa ttaaatgtac
     5941 attttccttt cttcgcccgt tgataagcga acgcgatgtg gcgcaggcaa tgtgttgctc
     6001 ttgcgacaca aacgcaatca aaatggattc aatttcgctt tttcccagtg aaacgaagaa
     6061 cgaaccgacc atcatgatat gctcctctgc atgttgcgta ttgaatcaat gacaatttca
     6121 attaagccgc ccgttcgtca tgcgttttcg tgcgcttcga aatgctgata acgctgctgt
     6181 cctccaactg ctttgcatgt ggacacaatt ccatttattt aattctttta tttggatcgg
     6241 ttaaattaaa aagcgccttg ttacgcattt aacgttgttt ccggtgcgtg gtggtttcat
     6301 gcttctggga acggcaaatg ggtttaggat tgggaacccc tcatcatctg ttggaatata
     6361 ctattcaacc tacaaaagta acgttaaaca acactacttt atatttgata tgaatggcca
     6421 caccttttat gccataaaac atattgtaag agaataccac tctttttatt ccttctttcc
     6481 ttcttgtacg ttttttgctg taagtaggtc gtggtgctgg tgttgcagtt gaaataactt
     6541 aaaatataaa tcataaaact caaacataaa cttgactatt tatttattta ttaagaaagg
     6601 aaatataaat tataaattac aacaggttat ggacctgcag ccaagcttgg cgctcgtccg
     6661 ggggcaatga gatatgaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat
     6721 cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc
     6781 tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg
     6841 tttctacaaa gatcgttatg tttatcggca ctttgcatcg gccgcgctcc cgattccgga
     6901 agtgcttgac attggggaat tcagcgagag cctgacctat tgcatctccc gccgtgcaca
     6961 gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc
     7021 ggaggccatg gatgcgatcg ctgcggccga tcttagccag acgagcgggt tcggcccatt
     7081 cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga
     7141 tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca
     7201 ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca
     7261 cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga
     7321 ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag
     7381 gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct
     7441 tgcaggatcg ccgcggctcc gggcgtatat gccccgcatt ggtcttgacc aactctatca
     7501 gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat
     7561 cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt
     7621 ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg
     7681 tccgagggca aaggaataga gtagatgccg accgaacaag agctgatttc gagaacgcct
     7741 cagccagcaa ctcgc
//
