LOCUS       sequence-412943-        6507 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression plasmid for Gal4DBD-HIF-CAD.myc expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6507)
  AUTHORS   Allen TP, Roennfeldt AE, Reckdharajkumar M, Liu M, Quinn RJ, Russell
            DL, Peet DJ, Whitelaw ML, Bersten DC
  TITLE     dFLASH; dual FLuorescent transcription factor Activity Sensor for 
            Histone integrated live-cell reporting and high-content screening
  JOURNAL   bioRxiv 2023.11.21.568191
REFERENCE   2  (bases 1 to 6507)
  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: "bioRxiv 
            2023.11.21.568191"
COMMENT     Sequence Label: pEF-IRES-puro6 gal4DBD-HIFCAD myc tag
FEATURES             Location/Qualifiers
     source          1..6507
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        16..1178
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          247..1169
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1126..1146
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     intron          1217..1349
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1332..1351
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1394..1412
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1478..1918
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     CDS             1937..1966
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     misc_feature    2314..2866
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2458..2475)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2685..2704
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2866..3468
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MGTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     3365..3385
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3705..3826
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3742..3761)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3796..3815
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3917..4372
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4004..4023)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4214..4235
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4441..4460)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4560..4582
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4620..4638)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4706..4810
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4811..4879,4880..5671)
                     /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     complement(5029..5048)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5842..6430
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6331..6350
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 attcaagctt cgtgaggctc cggtgcccgt cagtgggcag agcgcacatc gcccacagtc
       61 cccgagaagt tggggggagg ggtcggcaat tgaaccggtg cctagagaag gtggcgcggg
      121 gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa
      181 ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt tgccgccaga
      241 acacaggtaa gtgccgtgtg tggttcccgc gggcctggcc tctttacggg ttatggccct
      301 tgcgtgcctt gaattacttc cacctggctc cagtacgtga ttcttgatcc cgagctggag
      361 ccaggggcgg gccttgcgct ttaggagccc cttcgcctcg tgcttgagtt gaggcctggc
      421 ctgggcgctg gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt
      481 tcgataagtc tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc
      541 aagatagtct tgtaaatgcg ggccaggatc tgcacactgg tatttcggtt tttggggccg
      601 cgggcggcga cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag
      661 cgcggccacc gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg
      721 gcctcgcgcc gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag
      781 ttgcgtgagc ggaaagatgg ccgcttcccg gccctgctcc agggggctca aaatggagga
      841 cgcggcgctc gggagagcgg gcgggtgagt cacccacaca aaggaaaggg gcctttccgt
      901 cctcagccgt cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt
      961 agttctggag cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg
     1021 agtttcccca cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat
     1081 tctccttgga atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag
     1141 tggttcaaag tttttttctt ccatttcagg tgtcgtgaac acgggctgca gaagttggtc
     1201 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
     1261 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
     1321 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1381 aggctagagt acttaatacg actcactata ggctagcctc gagaattcac gcgtggtacc
     1441 tctagagatc tcatatggat ccactagtat cgatgatatg aagctactgt cttctatcga
     1501 acaagcatgc gatatttgcc gacttaaaaa gctcaagtgc tccaaagaaa aaccgaagtg
     1561 cgccaagtgt ctgaagaaca actgggagtg tcgctactct cccaaaacca aaaggtctcc
     1621 gctgactagg gcacatctga cagaagtgga atcaaggcta gaaagactgg aacagctatt
     1681 tctactgatt tttcctcgag aagaccttga catgattttg aaaatggatt ctttacagga
     1741 tataaaagca ttgttaacag gattatttgt acaagataat gtgaataaag atgccgtcac
     1801 agatagattg gcttcagtgg agactgatat gcctctaaca ttgagacagc atagaataag
     1861 tgcgacatca tcatcggaag agagtagtaa caaaggtcaa agacagttga ctgtatcgga
     1921 attcgttatc cgaccagaac aaaagttgat ttctgaagaa gatttgtcac tttttcaagc
     1981 agtaggaatt ggaacattat tacagcagcc agacgatcat gcagctacta catcactttc
     2041 ttggaaacgt gtaaaaggat gcaaatctag tgaacagaat ggaatggagc aaaagacaat
     2101 tattttaata ccctctgatt tagcatgtag actgctgggg caatcaatgg atgaaagtgg
     2161 attaccacag ctgaccagtt atgattgtga agttaatgct cctatacaag gcagcagaaa
     2221 cttactgcag ggtgaagaat tactcagagc tttggatcaa gttaactgaa tcgagctcgc
     2281 ggccgctcta gcccaattcc gcccctcccc ctaacgttac tggccgaagc cgcttggaat
     2341 aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct tttggcaatg
     2401 tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt ctttcccctc
     2461 tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct ctggaagctt
     2521 cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc ccacctggcg
     2581 acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag gcggcacaac
     2641 cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc tcctcaagcg
     2701 tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga tctgatctgg
     2761 ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc
     2821 cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataatatggg caccgagtac
     2881 aagcccacgg tgcgcctcgc cacccgcgac gacgtccccc gggccgtacg caccctcgcc
     2941 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg acccggaccg ccacatcgag
     3001 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     3061 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     3121 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     3181 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     3241 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     3301 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     3361 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     3421 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac
     3481 gacccgcagc gcccgaccga aaggagcgca cgaccccatg atggctccga ccgaagccac
     3541 ccggggcggc cccgccgacc ccgcacccgc ccccgaggcc caccgactct agctagagga
     3601 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     3661 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     3721 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     3781 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatga
     3841 tccgggctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
     3901 cctgaatggc gaatggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     3961 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     4021 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     4081 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     4141 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     4201 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     4261 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     4321 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttcctgatgc
     4381 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatatggt gcactctcag
     4441 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     4501 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     4561 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     4621 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
     4681 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     4741 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     4801 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     4861 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     4921 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     4981 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     5041 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     5101 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     5161 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     5221 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     5281 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     5341 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     5401 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     5461 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     5521 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     5581 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     5641 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     5701 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     5761 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     5821 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     5881 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5941 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     6001 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     6061 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     6121 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     6181 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     6241 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     6301 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     6361 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     6421 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     6481 tgctcacatg gctcgacaga tccccca
//
