LOCUS       sequence-129526-        6697 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  flySTARR-seq validation vector with Gateway cassette downstream of 
            the firefly luciferase gene and the RpS12 core promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6697)
  AUTHORS   Zabidi MA, Arnold CD, Schernhuber K, Pagani M, Rath M, Frank O, 
            Stark A
  TITLE     Enhancer-core-promoter specificity separates developmental and 
            housekeeping gene regulation.
  JOURNAL   Nature. 2015 Feb 26;518(7540):556-9. doi: 10.1038/nature13994. Epub 
            2014 Dec 15.
  PUBMED    25517091
REFERENCE   2  (bases 1 to 6697)
  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: 
            "10.1038/nature13994"; journalName: "Nature"; date: "2015-02-26- 
            26"; volume: "518"; issue: "7540"; pages: "556-9"
COMMENT     Sequence Label: pGL3-RpS12-Gateway
FEATURES             Location/Qualifiers
     source          1..6697
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    7..55
                     /note="synthetic polyadenylation signal"
     misc_feature    69..160
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     109..128
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     CDS             429..2081
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(495..513)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    2122..2243
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2133..2152)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2187..2206
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    2478..2602
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2627..2632,2633..2650,2651..2657)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2711..3370
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2777..2796)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3712..4017
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3896..3915
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4058..4182)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4196..4213)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4367..4955)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4447..4466)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5126..5917,5918..5986))
                     /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     5749..5768
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5987..6091)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6118..6573
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6205..6224)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6415..6436
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ggccgcaata aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg
       61 atagtactaa catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag
      121 gctgtcccca gtgcaagtgc aggtgccaga acatttctct atcgataggt accgagctct
      181 tacgcgtcca tgcatgctat gggaattccc atcagacgtc tggctcgaga tctgcaatct
      241 tgtaccaata gctaaaaact cacatctcca gcgccatgcc gattttgttc tctttctttc
      301 cggttgtcaa aaggtacaga tgcttggatt ttatttctcc gaaatgaaga ggttttctta
      361 tcgaaaatgt aataaatatg aacaattaac tatcttttcc agtgcagtgc atccttaacc
      421 gcagaacaat ggaagacgcc aaaaacataa agaaaggccc ggcgccattc tatccgctgg
      481 aagatggaac cgctggagag caactgcata aggctatgaa gagatacgcc ctggttcctg
      541 gaacaattgc ttttacagat gcacatatcg aggtggacat cacttacgct gagtacttcg
      601 aaatgtccgt tcggttggca gaagctatga aacgatatgg gctgaataca aatcacagaa
      661 tcgtcgtatg cagtgaaaac tctcttcaat tctttatgcc ggtgttgggc gcgttattta
      721 tcggagttgc agttgcgccc gcgaacgaca tttataatga acgtgaattg ctcaacagta
      781 tgggcatttc gcagcctacc gtggtgttcg tttccaaaaa ggggttgcaa aaaattttga
      841 acgtgcaaaa aaagctccca atcatccaaa aaattattat catggattct aaaacggatt
      901 accagggatt tcagtcgatg tacacgttcg tcacatctca tctacctccc ggttttaatg
      961 aatacgattt tgtgccagag tccttcgata gggacaagac aattgcactg atcatgaact
     1021 cctctggatc tactggtctg cctaaaggtg tcgctctgcc tcatagaact gcctgcgtga
     1081 gattctcgca tgccagagat cctatttttg gcaatcaaat cattccggat actgcgattt
     1141 taagtgttgt tccattccat cacggttttg gaatgtttac tacactcgga tatttgatat
     1201 gtggatttcg agtcgtctta atgtatagat ttgaagaaga gctgtttctg aggagccttc
     1261 aggattacaa gattcaaagt gcgctgctgg tgccaaccct attctccttc ttcgccaaaa
     1321 gcactctgat tgacaaatac gatttatcta atttacacga aattgcttct ggtggcgctc
     1381 ccctctctaa ggaagtcggg gaagcggttg ccaagaggtt ccatctgcca ggtatcaggc
     1441 aaggatatgg gctcactgag actacatcag ctattctgat tacacccgag ggggatgata
     1501 aaccgggcgc ggtcggtaaa gttgttccat tttttgaagc gaaggttgtg gatctggata
     1561 ccgggaaaac gctgggcgtt aatcaaagag gcgaactgtg tgtgagaggt cctatgatta
     1621 tgtccggtta tgtaaacaat ccggaagcga ccaacgcctt gattgacaag gatggatggc
     1681 tacattctgg agacatagct tactgggacg aagacgaaca cttcttcatc gttgaccgcc
     1741 tgaagtctct gattaagtac aaaggctatc aggtggctcc cgctgaattg gaatccatct
     1801 tgctccaaca ccccaacatc ttcgacgcag gtgtcgcagg tcttcccgac gatgacgccg
     1861 gtgaacttcc cgccgccgtt gttgttttgg agcacggaaa gacgatgacg gaaaaagaga
     1921 tcgtggatta cgtcgccagt caagtaacaa ccgcgaaaaa gttgcgcgga ggagttgtgt
     1981 ttgtggacga agtaccgaaa ggtcttaccg gaaaactcga cgcaagaaaa atcagagaga
     2041 tcctcataaa ggccaagaag ggcggaaaga tcgccgtgta attctagagt cggggcggcc
     2101 ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     2161 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     2221 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     2281 agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcg
     2341 ataaggatcc gtcgaccgat gcccttgaga gccttcaacc cagtcagctc cttccggtgg
     2401 gcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat gcaactcgta
     2461 ggacaggtgc cggcagcaca agtttgtaca aaaaagctga acgagaaacg taaaatgata
     2521 taaatatcaa tatattaaat tagattttgc ataaaaaaca gactacataa tactgtaaaa
     2581 cacaacatat ccagtcacta tggcggccgc attaggcacc ccaggcttta cactttatgc
     2641 ttccggctcg tataatgtgt ggattttgag ttaggatccg tcgagatttt caggagctaa
     2701 ggaagctaaa atggagaaaa aaatcactgg atataccacc gttgatatat cccaatggca
     2761 tcgtaaagaa cattttgagg catttcagtc agttgctcaa tgtacctata accagaccgt
     2821 tcagctggat attacggcct ttttaaagac cgtaaagaaa aataagcaca agttttatcc
     2881 ggcctttatt cacattcttg cccgcctgat gaatgctcat ccggaattcc gtatggcaat
     2941 gaaagacggt gagctggtga tatgggatag tgttcaccct tgttacaccg ttttccatga
     3001 gcaaactgaa acgttttcat cgctctggag tgaataccac gacgatttcc ggcagtttct
     3061 acacatatat tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg
     3121 gtttattgag aatatgtttt tcgtctcagc caatccctgg gtgagtttca ccagttttga
     3181 tttaaacgtg gccaatatgg acaacttctt cgcccccgtt ttcaccatgg gcaaatatta
     3241 tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga
     3301 tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg
     3361 cggggcgtaa acgcgtggat ccggcttact aaaagccaga taacagtatg cgtatttgcg
     3421 cgctgatttt tgcggtataa gaatatatac tgatatgtat acccgaagta tgtcaaaaag
     3481 aggtatgcta tgaagcagcg tattacagtg acagttgaca gcgacagcta tcagttgctc
     3541 aaggcatata tgatgtcaat atctccggtc tggtaagcac aaccatgcag aatgaagccc
     3601 gtcgtctgcg tgccgaacgc tggaaagcgg aaaatcagga agggatggct gaggtcgccc
     3661 ggtttattga aatgaacggc tcttttgctg acgagaacag gggctggtga aatgcagttt
     3721 aaggtttaca cctataaaag agagagccgt tatcgtctgt ttgtggatgt acagagtgat
     3781 attattgaca cgcccgggcg acggatggtg atccccctgg ccagtgcacg tctgctgtca
     3841 gataaagtct cccgtgaact ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg
     3901 atgaccaccg atatggccag tgtgccggtc tccgttatcg gggaagaagt ggctgatctc
     3961 agccaccgcg aaaatgacat caaaaacgcc attaacctga tgttctgggg aatataaatg
     4021 tcaggctccc ttatacacag ccagtctgca ggtcgaccat agtgactgga tatgttgtgt
     4081 tttacagtat tatgtagtct gttttttatg caaaatctaa tttaatatat tgatatttat
     4141 atcattttac gtttctcgtt cagctttctt gtacaaagtg gtgctcttcc gcttcctcgc
     4201 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     4261 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4321 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4381 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4441 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     4501 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4561 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4621 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4681 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4741 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4801 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4861 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4921 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4981 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5041 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     5101 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5161 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5221 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     5281 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5341 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     5401 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     5461 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     5521 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     5581 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     5641 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     5701 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     5761 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     5821 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     5881 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     5941 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6001 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6061 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     6121 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     6181 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     6241 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     6301 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     6361 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     6421 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     6481 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     6541 cgaattttaa caaaatatta acgcttacaa tttgccattc gccattcagg ctgcgcaact
     6601 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagcccaag ctaccatgat
     6661 aagtaagtaa tattaaggta cgggaggtac ttggagc
//
