LOCUS       sequence-318790-        6901 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of 3xFLAG-NLS-DsRed-Express2-ZF1 under the 
            EF1-alpha promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6901)
  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 6901)
  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: pJM505 EF1-alpha 3xFLAG-NLS-DsRed-Express2-ZF1 in 
            T...
FEATURES             Location/Qualifiers
     source          1..6901
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..66
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             73..93
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             136..810
                     /codon_start=1
                     /product="noncytotoxic tetrameric variant of DsRed 
                     fluorescent protein (Strack et al., 2008)"
                     /label=DsRed-Express2
                     /note="mammalian codon-optimized"
                     /translation="MDSTENVIKPFMRFKVHMEGSVNGHEFEIEGEGEGKPYEGTQTAK
                     LQVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGTFIYHVKFIGVNFPSDGPVMQKKTLGWEPSTERLYPRDGVLKGEIHK
                     ALKLKGGGHYLVEFKSIYMAKKPVKLPGYYYVDSKLDITSHNEDYTVVEQYERAEARHH
                     LFQ"
     primer_bind     complement(316..336)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     728..751
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(1167..1184)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1173..1397
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2166..2594
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2253..2272)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2463..2484
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2617..2813
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2664..2799
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2726..2745
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2933..3054
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2970..2989)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3024..3043
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3103..3119)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3103..3119)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3116..3138)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3127..3143
                     /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(3151..3157,3158..3175,3176..3181))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3196..3217
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3334..3351)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3505..4093)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3585..3604)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4264..5055,5056..5124))
                     /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     4887..4906
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5125..5229)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5610..6792
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          5841..6783
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     6740..6760
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     6794..6818
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      join(6893..6901,1)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 gattataagg atcacgatgg cgactacaag gaccacgata ttgactacaa agacgatgac
       61 gataaaatgg ctcctaagaa aaagcgcaaa gtcggtatcc atggcgttcc cggatcaggc
      121 tccggaagtg gctccatgga tagcactgag aacgtcatca agcccttcat gcgcttcaag
      181 gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc
      241 aagccctacg agggcaccca gaccgccaag ctccaggtga ccaagggcgg ccccctgccc
      301 ttcgcctggg acatcctgtc cccccagttc cagtacggct ccaaggtgta cgtgaagcac
      361 cccgccgaca tccccgacta caagaagctg tccttccccg agggcttcaa gtgggagcgc
      421 gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctccaggac
      481 ggcaccttca tctaccacgt gaagttcatc ggcgtgaact tcccctccga cggccccgta
      541 atgcagaaga aaactctggg ctgggagccc tccaccgagc gcctgtaccc ccgcgacggc
      601 gtgctgaagg gcgagatcca caaggcgctg aagctgaagg gcggcggcca ctacctggtg
      661 gagttcaagt caatctacat ggccaagaag cccgtgaagc tgcccggcta ctactacgtg
      721 gactccaagc tggacatcac ctcccacaac gaggactaca ccgtggtgga gcagtacgag
      781 cgcgccgagg cccgccacca cctgttccag tctggttctg gttctggttc aggatccggt
      841 accggtggag ctagacccgg ggagcgcccc ttccagtgtc gcatttgcat gcggaacttt
      901 tcgcgccagg acaggcttga caggcatacc cgtactcata ccggtgaaaa accgtttcag
      961 tgtcggatct gtatgcgaaa tttctcccag aaggagcact tggcggggca tctacgtacg
     1021 cacaccggcg agaagccatt ccaatgccga atatgcatgc gcaacttcag tcgccgcgac
     1081 aacctgaacc ggcacctaaa aacccacctg agaggatctt aagcggccgc tcgagtctag
     1141 agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     1201 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     1261 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     1321 tggggtgggg caggacagca agggggagga ttgggaagtc aatagcaggc atgctgggga
     1381 tgcggtgggc tctatggctt ctgaggcagg tcgaagaact atttcctggc tgttacgcga
     1441 gcgcatcgcg ataagctcta gagggacagc ccccccccaa agcccccagg gatgtaatta
     1501 cgtccctccc ccgctagggg gcagcagcga gccgcccggg gctccgctcc ggtccggcgc
     1561 tccccccgca tccccgagcc ggcagcgtgc ggggacagcc cgggcacggg gaaggtggca
     1621 cgggatcgct ttcctctgaa cgcttctcgc tgctctttga gcctgcagac acctgggggg
     1681 atacggggaa aaagctttag gctgaaagag agatttagaa tgacagaatc atagaacggc
     1741 ctgcgtacgg atggggctct agctctagag cttgatatcg aattcctgca gccccgctag
     1801 agggacagcc cccccccaaa gcccccaggg atgtaattac gtccctcccc cgctaggggg
     1861 cagcagcgag ccgcccgggg ctccgctccg gtccggcgct ccccccgcat ccccgagccg
     1921 gcagcgtgcg gggacagccc gggcacgggg aaggtggcac gggatcgctt tcctctgaac
     1981 gcttctcgct gctctttgag cctgcagaca cctgggggga tacggggaaa aagctttagg
     2041 ctgaaagaga gatttagaat gacagaatca tagaacggcc tgggcgcgcc tggtggcgaa
     2101 cgatcaacag acgggagagg actaaagtct tcgctgacag cgaccggact ctagggggta
     2161 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2221 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     2281 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     2341 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     2401 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     2461 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     2521 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     2581 atttaacgcg aattaattct gtggaatgtg tgtcagttag gctcaattag tcagcaacca
     2641 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     2701 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     2761 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     2821 cgggagcttg tatatccatt ttcggaggtt gggcttcgga atcgttttcc gggacgccgg
     2881 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     2941 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     3001 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     3061 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     3121 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     3181 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     3241 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     3301 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     3361 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     3421 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     3481 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     3541 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     3601 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     3661 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     3721 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     3781 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     3841 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     3901 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     3961 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4021 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4081 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     4141 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     4201 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     4261 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     4321 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     4381 ccccagtgct gcaatgatac cgcgagatcc acgctcaccg gctccagatt tatcagcaat
     4441 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     4501 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     4561 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     4621 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     4681 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     4741 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     4801 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     4861 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     4921 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     4981 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5041 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     5101 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     5161 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     5221 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatcgaag
     5281 acgggcaaaa ccgatgttga cggactaatc ctgacaagat ctggggaggt cgctgagtag
     5341 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
     5401 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgtgag
     5461 taattcatac aaaaggactc gcccctgcct tggggaatcc cagggaccgt cgttaaactc
     5521 ccactaacgt agaacccaga gatcgctgcg ttcccgcccc ctcacccgcc cgctctcgtc
     5581 atcactgagg tggagaagag catgcgtgag gctccggtgc ccgtcagtgg gcagagcgca
     5641 catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc ggtgcctaga
     5701 gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg
     5761 agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg
     5821 ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct ggcctcttta
     5881 cgggttatgg cccttgcgtg ccttgaatta cttccacgcc cctggctgca gtacgtgatt
     5941 cttgatcccg agcttcgggt tggaagtggg tgggagagtt cgaggccttg cgcttaagga
     6001 gccccttcgc ctcgtgcttg agttgaggcc tggcttgggc gctggggccg ccgcgtgcga
     6061 atctggtggc accttcgcgc ctgtctcgct gctttcgata agtctctagc catttaaaat
     6121 ttttgatgac ctgctgcgac gctttttttc tggcaagata gtcttgtaaa tgcgggccaa
     6181 gatctgcaca ctggtatttc ggtttttggg gccgcgggcg gcgacggggc ccgtgcgtcc
     6241 cagcgcacat gttcggcgag gcggggcctg cgagcgcggc caccgagaat cggacggggg
     6301 tagtctcaag ctggccggcc tgctctggtg cctggcctcg cgccgccgtg tatcgccccg
     6361 ccctgggcgg caaggctggc ccggtcggca ccagttgcgt gagcggaaag atggccgctt
     6421 cccggccctg ctgcagggag ctcaaaatgg aggacgcggc gctcgggaga gcgggcgggt
     6481 gagtcaccca cacaaaggaa aagggccttt ccgtcctcag ccgtcgcttc atgtgactcc
     6541 acggagtacc gggcgccgtc caggcacctc gattagttct cgagcttttg gagtacgtcg
     6601 tctttaggtt ggggggaggg gttttatgcg atggagtttc cccacactga gtgggtggag
     6661 actgaagtta ggccagcttg gcacttgatg taattctcct tggaatttgc cctttttgag
     6721 tttggatctt ggttcattct caagcctcag acagtggttc aaagtttttt tcttccattt
     6781 caggtgtcgt gagagctcgt ttagtgaacc gtcagatcgc ctggagacgc catccacgct
     6841 gttttgacct ccatagaaga gaacgggacc gatccaaatt ctcgaggcta gcgccaccat
     6901 g
//
