LOCUS       sequence-412944-        6483 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression plasmid for Gal4DBD-HIF-CAD  expression GalO linker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6483)
  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 6483)
  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 +gal4DBDHIFCAD pGalO linker
FEATURES             Location/Qualifiers
     source          1..6483
                     /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"
     misc_feature    2274..2840
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2290..2842
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2434..2451)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2661..2680
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2842..3444
                     /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     3341..3361
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3681..3802
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3718..3737)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3772..3791
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3893..4348
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3980..3999)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4190..4211
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4417..4436)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4536..4558
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4596..4614)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4682..4786
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4787..4855,4856..5647)
                     /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(5005..5024)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5818..6406
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6307..6326
                     /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 ctgtatcgcc
     1921 ggaattcccg gggatccgtc gatcactttt tcaagcagta ggaattggaa cattattaca
     1981 gcagccagac gatcatgcag ctactacatc actttcttgg aaacgtgtaa aaggatgcaa
     2041 atctagtgaa cagaatggaa tggagcaaaa gacaattatt ttaataccct ctgatttagc
     2101 atgtagactg ctggggcaat caatggatga aagtggatta ccacagctga ccagttatga
     2161 ttgtgaagtt aatgctccta tacaaggcag cagaaactta ctgcagggtg aagaattact
     2221 cagagctttg gatcaagtta actgaatcga gctcgcggcc gctctagccc aattccgccc
     2281 ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg tttgtctata
     2341 tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg
     2401 tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg caaggtctgt
     2461 tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca acgtctgtag
     2521 cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc ggccaaaagc
     2581 cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga
     2641 tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg ctgaaggatg
     2701 cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca tgctttacat
     2761 gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac gtggttttcc
     2821 tttgaaaaac acgatgataa tatgggcacc gagtacaagc ccacggtgcg cctcgccacc
     2881 cgcgacgacg tcccccgggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     2941 acgcgccaca ccgtcgaccc ggaccgccac atcgagcggg tcaccgagct gcaagaactc
     3001 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg
     3061 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     3121 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc
     3181 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     3241 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag
     3301 cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag
     3361 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     3421 atgacccgca agcccggtgc ctgacgcccg ccccacgacc cgcagcgccc gaccgaaagg
     3481 agcgcacgac cccatgatgg ctccgaccga agccacccgg ggcggccccg ccgaccccgc
     3541 acccgccccc gaggcccacc gactctagct agaggatcat aatcagccat accacatttg
     3601 tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa
     3661 tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca
     3721 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     3781 ccaaactcat caatgtatct tatcatgtct ggatgatccg ggctggcgta atagcgaaga
     3841 ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggacgcgccc
     3901 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     3961 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4021 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4081 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     4141 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     4201 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4261 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4321 tttaacaaaa tattaacgct tacaatttcc tgatgcggta ttttctcctt acgcatctgt
     4381 gcggtatttc acaccgcata tatggtgcac tctcagtaca atctgctctg atgccgcata
     4441 gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     4501 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     4561 ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata
     4621 ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt
     4681 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     4741 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     4801 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     4861 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     4921 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     4981 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     5041 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     5101 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     5161 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     5221 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     5281 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     5341 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     5401 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     5461 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     5521 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     5581 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     5641 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     5701 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     5761 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5821 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5881 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5941 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     6001 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     6061 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     6121 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6181 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     6241 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     6301 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     6361 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     6421 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatggctc gacagatccc
     6481 cca
//
