LOCUS       sequence-317637-        6877 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of TC 
            3xFLAG-FRB-FGFR4-PRS(M)-(GS)-NLS-VP64-ZF1 under the EF1-alpha 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6877)
  AUTHORS   Muldoon JJ, Kandula V, Hong M, Donahue PS, Boucher JD, Bagheri N, 
            Leonard JN
  TITLE     Model-guided design of mammalian genetic programs.
  JOURNAL   Sci Adv. 2021 Feb 19;7(8). pii: 7/8/eabe9375. doi: 
            10.1126/sciadv.abe9375. Print 2021 Feb.
  PUBMED    33608279
REFERENCE   2  (bases 1 to 6877)
  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: "Sci Adv."; 
            date: "2021-02-19"; volume: "7(8). pii: 7/8/eabe9375. doi"; pages: "
            10.1126/sciadv.abe9375. Print 2021 Feb"
COMMENT     Sequence Label: pJM612 EF1-alpha TC
            3xFLAG-FRB-FGFR4-PRS(M)-(GS)-NL...
FEATURES             Location/Qualifiers
     source          1..6877
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          231..1173
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1130..1150
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1184..1208
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1367..1432
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1832..1852
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             1859..1879
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1904..2053
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     primer_bind     complement(2434..2451)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2440..2664
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3433..3861
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3520..3539)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3730..3751
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3884..4080
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3931..4066
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3993..4012
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    4200..4321
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4237..4256)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4291..4310
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4370..4386)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4370..4386)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4383..4405)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4394..4410
                     /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(4418..4424,4425..4442,4443..4448))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4463..4484
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4601..4618)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4772..5360)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4852..4871)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5531..6322,6323..6391))
                     /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     6154..6173
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6392..6496)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(6877,1..1182)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
ORIGIN
        1 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
       61 gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
      121 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
      181 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg
      241 tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta
      301 cttccacgcc cctggctgca gtacgtgatt cttgatcccg agcttcgggt tggaagtggg
      361 tgggagagtt cgaggccttg cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc
      421 tggcttgggc gctggggccg ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct
      481 gctttcgata agtctctagc catttaaaat ttttgatgac ctgctgcgac gctttttttc
      541 tggcaagata gtcttgtaaa tgcgggccaa gatctgcaca ctggtatttc ggtttttggg
      601 gccgcgggcg gcgacggggc ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg
      661 cgagcgcggc caccgagaat cggacggggg tagtctcaag ctggccggcc tgctctggtg
      721 cctggcctcg cgccgccgtg tatcgccccg ccctgggcgg caaggctggc ccggtcggca
      781 ccagttgcgt gagcggaaag atggccgctt cccggccctg ctgcagggag ctcaaaatgg
      841 aggacgcggc gctcgggaga gcgggcgggt gagtcaccca cacaaaggaa aagggccttt
      901 ccgtcctcag ccgtcgcttc atgtgactcc acggagtacc gggcgccgtc caggcacctc
      961 gattagttct cgagcttttg gagtacgtcg tctttaggtt ggggggaggg gttttatgcg
     1021 atggagtttc cccacactga gtgggtggag actgaagtta ggccagcttg gcacttgatg
     1081 taattctcct tggaatttgc cctttttgag tttggatctt ggttcattct caagcctcag
     1141 acagtggttc aaagtttttt tcttccattt caggtgtcgt gagagctcgt ttagtgaacc
     1201 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga gaacgggacc
     1261 gatccaaatt ctcgaggcta gcgccatcat gtgccgagcc atctctctta ggcgcttgct
     1321 gctgctgctg ctgcagctgt cacaactcct agctgtcact caaggggatt acaaggatca
     1381 cgatggggat tacaaagatc atgacattga ctacaaggat gacgatgata agatgctcga
     1441 gatgtggcat gaaggcctgg aagaggcatc tcgtttgtac tttggggaaa ggaacgtgaa
     1501 aggcatgttt gaggtgctgg agcccttgca tgctatgatg gaacggggcc cccagactct
     1561 gaaggaaaca tcctttaatc aggcctatgg tcgagattta atggaggccc aagagtggtg
     1621 caggaagtac atgaaatcag ggaatgtcaa ggacctcctc caagcctggg acctctatta
     1681 tcatgtgttc cgacgaatct caaagactag aggaggatct ggaggctctg ggggttcagg
     1741 cggatcagga ggcagtggcg ggtcaaccgg tattatactg tatgctagtg gcagtctggc
     1801 attggcagtc ctcctgctgc tcgccgggtt ggagaacctg tattttcaga tgggatctcc
     1861 taagaaaaag cgcaaagtcg gtatccatgg cgttcccgga tcagacgcat tggacgattt
     1921 tgatctggat atgctgggaa gtgacgccct cgatgatttt gaccttgaca tgcttggttc
     1981 ggatgccctt gatgactttg acctcgacat gctcggcagt gacgcccttg atgatttcga
     2041 cctggacatg ctgattaact ctagaagttc cggatcttct ggttctggtt ctggttcagg
     2101 atccggtacc ggtggagcta gacccgggga gcgccccttc cagtgtcgca tttgcatgcg
     2161 gaacttttcg cgccaggaca ggcttgacag gcatacccgt actcataccg gtgaaaaacc
     2221 gtttcagtgt cggatctgta tgcgaaattt ctcccagaag gagcacttgg cggggcatct
     2281 acgtacgcac accggcgaga agccattcca atgccgaata tgcatgcgca acttcagtcg
     2341 ccgcgacaac ctgaaccggc acctaaaaac ccacctgaga ggatcttaag cggccgctcg
     2401 agtctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     2461 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     2521 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     2581 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagtcaat agcaggcatg
     2641 ctggggatgc ggtgggctct atggcttctg aggcaggtcg aagagactgc ccgacacatc
     2701 ttagtgagcg catcgcgata agctctagag ggacagcccc cccccaaagc ccccagggat
     2761 gtaattacgt ccctcccccg ctagggggca gcagcgagcc gcccggggct ccgctccggt
     2821 ccggcgctcc ccccgcatcc ccgagccggc agcgtgcggg gacagcccgg gcacggggaa
     2881 ggtggcacgg gatcgctttc ctctgaacgc ttctcgctgc tctttgagcc tgcagacacc
     2941 tggggggata cggggaaaaa gctttaggct gaaagagaga tttagaatga cagaatcata
     3001 gaacggcctg cgtacggatg gggctctagc tctagagctt gatatcgaat tcctgcagcc
     3061 ccgctagagg gacagccccc ccccaaagcc cccagggatg taattacgtc cctcccccgc
     3121 tagggggcag cagcgagccg cccggggctc cgctccggtc cggcgctccc cccgcatccc
     3181 cgagccggca gcgtgcgggg acagcccggg cacggggaag gtggcacggg atcgctttcc
     3241 tctgaacgct tctcgctgct ctttgagcct gcagacacct ggggggatac ggggaaaaag
     3301 ctttaggctg aaagagagat ttagaatgac agaatcatag aacggcctgg gcgcgcctgg
     3361 tggcgaacga tcaacagacg ggagaggtta caagtcttcg ctgacagcga ccggactcta
     3421 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     3481 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     3541 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     3601 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     3661 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     3721 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     3781 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     3841 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttaggct caattagtca
     3901 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     3961 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct
     4021 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     4081 aagctcccgg gagcttgtat atccattttc ggaggttggg cttcggaatc gttttccggg
     4141 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
     4201 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     4261 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     4321 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
     4381 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     4441 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     4501 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     4561 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4621 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4681 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4741 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4801 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4861 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4921 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4981 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     5041 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     5101 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     5161 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     5221 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5281 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5341 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5401 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5461 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5521 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5581 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5641 catctggccc cagtgctgca atgataccgc gagatccacg ctcaccggct ccagatttat
     5701 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5761 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5821 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5881 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5941 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6001 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6061 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6121 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6181 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6241 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6301 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6361 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6421 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6481 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag
     6541 atcgaagacg gactaaatag tagttcagac gccgttaagc gcaagatctg gggaggtcgc
     6601 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat
     6661 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
     6721 gcgtgagtaa ttcatacaaa aggactcgcc cctgccttgg ggaatcccag ggaccgtcgt
     6781 taaactccca ctaacgtaga acccagagat cgctgcgttc ccgccccctc acccgcccgc
     6841 tctcgtcatc actgaggtgg agaagagcat gcgtgag
//
