LOCUS       sequence-369216-        6693 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses EGFP tagged human 2N4R tau (WT) in mammalian cells with 
            CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6693)
  AUTHORS   Rajbanshi B, Guruacharya A, Mandell JW, Bloom GS
  TITLE     Localization, induction, and cellular effects of tau phosphorylated 
            at threonine 217.
  JOURNAL   Alzheimers Dement. 2023 Jan 12. doi: 10.1002/alz.12892.
  PUBMED    36633254
REFERENCE   2  (bases 1 to 6693)
  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: "Alzheimers 
            Dement. 2023 Jan 12. doi: 10.1002/alz.12892."
COMMENT     Sequence Label: EGFP-Tau(WT)
FEATURES             Location/Qualifiers
     source          1..6693
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     154..171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    288..309
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(324..329,330..347,348..354)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    362..378
                     /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)."
     primer_bind     367..389
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     386..402
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     386..402
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        431..810
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        811..1014
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     965..985
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1011..1035
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        1245..1263
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1245..1262
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1276..1295
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1339..1348
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1345..1347,1348..1350,1351..2061)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSNQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1390..1411)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1651..1670)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1998..2019
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      3410..3419
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3423..3557
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3460..3479)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3514..3533
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        3626..3983
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3626..3817
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3649..3669)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3834..3969
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4003..4020)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4003..4019)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4012..4034)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4232..4687
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4319..4338)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4529..4550
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4704..4723)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4823..4845
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4883..4901)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4969..5073
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5074..5142,5143..5934)
                     /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(5292..5311)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6105..6693
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6594..6613
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
       61 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
      121 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      181 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
      241 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
      301 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
      361 attgtgagcg gataacaatt tcacacagga aacagctatg acatgattac gaattaattc
      421 gagctcgccc gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
      481 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      541 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      601 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      661 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      721 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      781 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
      841 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
      901 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
      961 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta
     1021 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac
     1081 cgggaccgat ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc
     1141 aagagtgacg taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa
     1201 cgcggctaca attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat
     1261 agaataacat ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc
     1321 tcggttctat cgatggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg
     1381 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     1441 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     1501 aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc
     1561 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     1621 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     1681 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     1741 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     1801 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     1861 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     1921 cccgtgctgc tgcccgacaa ccactacctg agcaaccagt ccgccctgag caaagacccc
     1981 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     2041 ggcatggacg agctgtacaa gggatccgct gagccccgcc aggagttcga agtgatggaa
     2101 gatcacgctg ggacgtacgg gttgggggac aggaaagatc aggggggcta caccatgcac
     2161 caagaccaag agggtgacac ggacgctggc ctgaaagaat ctcccctgca gacccccact
     2221 gaggacggat ctgaggaacc gggctctgaa acctctgatg ctaagagcac tccaacagcg
     2281 gaagatgtga cagcaccctt agtggatgag ggagctcccg gcaagcaggc tgccgcgcag
     2341 ccccacacgg agatcccaga aggaaccaca gctgaagaag caggcattgg agacaccccc
     2401 agcctggaag acgaagctgc tggtcacgtg acccaagctc gcatggtcag taaaagcaaa
     2461 gacgggactg gaagcgatga caaaaaagcc aagggggctg atggtaaaac gaagatcgcc
     2521 acaccgcggg gagcagcccc tccaggccag aagggccagg ccaacgccac caggattcca
     2581 gcaaaaaccc cgcccgctcc aaagacacca cccagctctg gtgaacctcc aaaatcaggg
     2641 gatcgcagcg gctacagcag ccccggctcc ccaggcactc ccggcagccg ctcccgcacc
     2701 ccgtcccttc caaccccacc cacccgggag cccaagaagg tggcagtggt ccgtactcca
     2761 cccaagtcgc cgtcttccgc caagagccgc ctgcagacag cccccgtgcc catgccagac
     2821 ctgaagaatg tcaagtccaa gatcggctcc actgagaacc tgaagcacca gccgggaggc
     2881 gggaaggtgc agataattaa taagaagctg gatcttagca acgtccagtc caagtgtggc
     2941 tcaaaggata atatcaaaca cgtcccggga ggcggcagtg tgcaaatagt ctacaaacca
     3001 gttgacctga gcaaggtgac ctccaagtgt ggctcattag gcaacatcca tcataaacca
     3061 ggaggtggcc aggtggaagt aaaatctgag aagcttgact tcaaggacag agtccagtcg
     3121 aagattgggt ccctggacaa tatcacccac gtccctggcg gaggaaataa aaagattgaa
     3181 acccacaagc tgaccttccg cgagaacgcc aaagccaaga cagaccacgg ggcggagatc
     3241 gtgtacaagt cgccagtggt gtctggggac acgtctccac ggcatctcag caatgtctcc
     3301 tccaccggca gcatcgacat ggtagactcg ccccagctcg ccacgctagc tgacgaggtg
     3361 tctgcctccc tggccaagca gggtttgtga gtcgacctgc agaagcttgg ccgccatggc
     3421 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     3481 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     3541 cttatcatgt ctggatcggg aattaattcg gcgcagcacc atggcctgaa ataacctctg
     3601 aaagaggaac ttggttaggt accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc
     3661 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3721 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3781 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3841 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     3901 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     3961 ttggaggcct aggcttttgc aaaaagctgt taacagcttg gcactggccg tcgttttaca
     4021 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     4081 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     4141 cagcctgaat ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     4201 ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg
     4261 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     4321 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     4381 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     4441 cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     4501 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     4561 aaccctatct cgggctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     4621 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt
     4681 acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     4741 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     4801 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     4861 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     4921 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     4981 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     5041 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     5101 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     5161 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     5221 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     5281 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     5341 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     5401 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     5461 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     5521 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     5581 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     5641 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     5701 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     5761 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     5821 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     5881 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     5941 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     6001 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     6061 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     6121 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     6181 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     6241 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     6301 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     6361 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     6421 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     6481 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     6541 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     6601 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     6661 tgatgctcgt caggggggcg gagcctatgg aaa
//
