LOCUS       sequence-299333-        5444 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Flag tagged human FOXD4 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5444)
  AUTHORS   Moparthi L, Koch S
  TITLE     A uniform expression library for the exploration of FOX 
            transcription factor biology.
  JOURNAL   Differentiation. 2020 Sep - Oct;115:30-36. doi: 
            10.1016/j.diff.2020.08.002. Epub 2020 Aug 20.
  PUBMED    32858261
REFERENCE   2  (bases 1 to 5444)
  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: 
            "Differentiation. 2020 Sep - Oct"; volume: "115"; pages: "30-36. 
            doi: 10.1016/j.diff.2020.08.002. Epub 2020 Aug 20"
COMMENT     Sequence Label: Flag-FOXD4
FEATURES             Location/Qualifiers
     source          1..5444
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      85..94
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             94..117
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    1496..1630
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1520..1539)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1574..1593
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1743..1763)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1743..1761)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1782..1798)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1782..1798)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1795..1817)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1806..1822
                     /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)."
     promoter        complement(join(1830..1836,1837..1854,1855..1860))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1875..1896
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2013..2030)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2184..2772)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2264..2283)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2943..3734,3735..3803))
                     /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     3566..3585
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3804..3908)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3934..4389)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4071..4092)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4283..4302
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4460..5444
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atcagccacc atggattaca aggacgacga cgataaggga
      121 tcccatcgat tcgaattcaa cttgccaaga gctgagcgcc ttcgctccac accgcagcgc
      181 agcctccggg actccgatgg ggaagacggt aaaatcgatg tcttgggaga ggaggaagat
      241 gaagacgagg aggaggcggc gagccagcag ttcctagagc agtcgctcca gccggggctg
      301 caggtggccc ggtggggcgg ggttgcgctt ccccgagagc acatcgaggg cggcggcggc
      361 ccgagcgacc cctcagaatt tggcaccgag ttcagggcac cgccaaggtc tgcggcggcc
      421 tctgaagatg cccggcagcc ggcaaagccc ccctcctcgt acatcgcgct catcaccatg
      481 gccatcctgc aaagcccgca caagcgcctc acgctcagcg gcatctgcgc cttcattagt
      541 ggccgcttcc cctactaccg ccgcaagttc cccgcctggc agaacagcat ccgccacaac
      601 ctctcgctga acgactgctt cgtcaagatc ccccgcgagc cgggccgccc aggcaagggc
      661 aactactgga gcctggaccc cgcctcccag gacatgttcg acaatggcag ctttctccgg
      721 cgtaggaagc gtttccagcg ccaccaactg accccgggag cccacctgcc ccaccccttc
      781 cctctacctg ctgcacacgc cgccctgcac aacccccgcc caggccctct gcttggggcc
      841 cctgccccgc cgcagccagt cccgggggcc taccccaaca ccgcccccgg gagacgccct
      901 tacgctctgc tgcacccgca tcctcctcgc tacctactgc tctcggcccc cgcctatgcc
      961 ggggcaccga agaaagcaga aggcgcggac ctggcgaccc cggcaccctt cccgtgctgc
     1021 agccctcact tggtcctcag ccttgggagg agggcaaggg tctggcgtcg ccaccgggag
     1081 gcggatgcat ctctttcagc attgagagta tcatgcaagg ggtcagggga gcgggtacag
     1141 gggctgcgca gagtttgtcc ccgaccgcgt ggagctactg ccccctgctc cagcgaccgt
     1201 caagcctgtc ggacaatttt gcagcaacag cagcggcatc aggaggagga ctgcgccaac
     1261 ggctgcgctc ccaccaaggg cgcggtgctg ggcgggcacc tgtcggccgc gtcggcgctg
     1321 ctgcggtatc aggcggtggc agagggctct gggctgacat cgctggccgc ccctttgggc
     1381 ggagagggga cctcaccagt ttttttagta tcgcccacgc ccagttccct ggccgagtcc
     1441 gcagggccct cctagagcct cgagcctcta gaactatagt gagtcgtatt acgtagatcc
     1501 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     1561 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     1621 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     1681 ggaggttttt taattcgcgg ccgcggcgcc aatgcattgg gcccggtacc cagcttttgt
     1741 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     1801 tgaaattgtt atccgctcac aattccacac aacatacgag ccgggagcat aaagtgtaaa
     1861 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     1921 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     1981 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     2041 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     2101 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     2161 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2221 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2281 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2341 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     2401 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     2461 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     2521 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     2581 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     2641 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     2701 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     2761 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     2821 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     2881 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     2941 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     3001 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     3061 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     3121 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     3181 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     3241 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     3301 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     3361 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     3421 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     3481 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     3541 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     3601 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     3661 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     3721 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     3781 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     3841 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     3901 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     3961 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     4021 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     4081 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     4141 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     4201 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     4261 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     4321 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     4381 agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     4441 cctcttcgct attacgccag tcgaccgcca attcaatatg gcgtatatgg actcatgcca
     4501 attcaatatg gtggatctgg acctgtgcca attcaatatg gcgtatatgg actcgtgcca
     4561 attcaatatg gtggatctgg accccagcca attcaatatg gcggacttgg caccatgcca
     4621 attcaatatg gcggacctgg cactgtgcca actggggagg ggtctacttg gcacggtgcc
     4681 aagtttgagg aggggtcttg gccctgtgcc aagtccgcca tattgaattg gcatggtgcc
     4741 aataatggcg gccatattgg ctatatgcca ggatcaatat ataggcaata tccaatatgg
     4801 ccctatgcca atatggctat tggccaggtt caatactatg tattggccct atgccatata
     4861 gtattccata tatgggtttt cctattgacg tagatagccc ctcccaatgg gcggtcccat
     4921 ataccatata tggggcttcc taataccgcc catagccact cccccattga cgtcaatggt
     4981 ctctatatat ggtctttcct attgacgtca tatgggcggt cctattgacg tatatggcgc
     5041 ctcccccatt gacgtcaatt acggtaaatg gcccgcctgg ctcaatgccc attgacgtca
     5101 ataggaccac ccaccattga cgtcaatggg atggctcatt gcccattcat atccgttctc
     5161 acgcccccta ttgacgtcaa tgacggtaaa tggcccactt ggcagtacat caatatctat
     5221 taatagtaac ttggcaagta cattactatt ggaagtacgc cagggtacat tggcagtact
     5281 cccattgacg tcaatggcgg taaatggccc gcgatggctg ccaagtacat ccccattgac
     5341 gtcaatgggg aggggcaatg acgcaaatgg gcgttccatt gacgtaaatg ggcggtaggc
     5401 gtgcctaatg ggaggtctat ataagcaatg ctcgtttagg gaac
//
