LOCUS       sequence-48237-e        5963 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5963)
  AUTHORS   Roczniak-Ferguson A, Petit CS, Froehlich F, Qian S, Ky J, Angarola 
            B, Walther TC, Ferguson SM
  TITLE     The Transcription Factor TFEB Links mTORC1 Signaling to 
            Transcriptional Control of Lysosome Homeostasis.
  JOURNAL   Sci Signal. 2012 Jun 12;5(228):ra42.
  PUBMED    22692423
REFERENCE   2  (bases 1 to 5963)
  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 
            Signal."; date: "2012-06-12"; volume: "5(228)"; pages: "ra42"
COMMENT     Sequence Label: pEGFP-N1-MITF-M
FEATURES             Location/Qualifiers
     source          1..5963
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      1903..1912
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1909..1911,1912..1914,1915..2628)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1909..1911,1912..1914,1915..2625)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1954..1975)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2215..2234)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2562..2583
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2751..2872
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2788..2807)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2842..2861
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2879..3334)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3016..3037)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3228..3247
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3361..3465
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3467..3824
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3467..3658
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3490..3510)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3675..3810
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3859..4653
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3859..4653
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3913..3932)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4523..4542
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4841..4860)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4885..4932
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5261..5849
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5750..5769
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttgac accatgctgg aaatgctaga atataatcac
      661 tatcaggtgc agacccacct cgaaaacccc accaagtacc acatacagca agcccaacgg
      721 cagcaggtaa agcagtacct ttctaccact ttagcaaata aacatgccaa ccaagtcctg
      781 agcttgccat gtccaaacca gcctggcgat catgtcatgc caccggtgcc ggggagcagc
      841 gcacccaaca gccccatggc tatgcttacg cttaactcca actgtgaaaa agagggattt
      901 tataagtttg aagagcaaaa cagggcagag agcgagtgcc caggcatgaa cacacattca
      961 cgagcgtcct gtatgcagat ggatgatgta atcgatgaca tcattagcct agaatcaagt
     1021 tataatgagg aaatcttggg cttgatggat cctgctttgc aaatggcaaa tacgttgcct
     1081 gtctcgggaa acttgattga tctttatgga aaccaaggtc tgcccccacc aggcctcacc
     1141 atcagcaact cctgtccagc caaccttccc aacataaaaa gggagctcac agagtctgaa
     1201 gcaagagcac tggccaaaga gaggcagaaa aaggacaatc acaacctgat tgaacgaaga
     1261 agaagattta acataaatga ccgcattaaa gaactaggta ctttgattcc caagtcaaat
     1321 gatccagaca tgcgctggaa caagggaacc atcttaaaag catccgtgga ctatatccga
     1381 aagttgcaac gagaacagca acgcgcaaaa gaacttgaaa accgacagaa gaaactggag
     1441 cacgccaacc ggcatttgtt gctcagaata caggaacttg aaatgcaggc tcgagctcat
     1501 ggactttccc ttattccatc cacgggtctc tgctctccag atttggtgaa tcggatcatc
     1561 aagcaagaac ccgttcttga gaactgcagc caagacctcc ttcagcatca tgcagaccta
     1621 acctgtacaa caactctcga tctcacggat ggcaccatca ccttcaacaa caacctcgga
     1681 actgggactg aggccaacca agcctatagt gtccccacaa aaatgggatc caaactggaa
     1741 gacatcctga tggacgacac cctttctccc gtcggtgtca ctgatccact cctttcctca
     1801 gtgtcccccg gagcttccaa aacaagcagc cggaggagca gtatgagcat ggaagagacg
     1861 gagcacactt gtacggtacc gcgggcccgg gatccaccgg tcgccaccat ggtgagcaag
     1921 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     1981 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     2041 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     2101 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     2161 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     2221 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     2281 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     2341 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     2401 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     2461 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     2521 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     2581 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaaag cggccgcgac
     2641 tctagatcat aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc
     2701 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta
     2761 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     2821 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct taaggcgtaa
     2881 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     2941 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     3001 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     3061 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     3121 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     3181 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     3241 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     3301 cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg
     3361 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     3421 caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga
     3481 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     3541 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     3601 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     3661 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3721 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     3781 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca
     3841 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     3901 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     3961 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     4021 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
     4081 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     4141 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     4201 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     4261 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     4321 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     4381 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     4441 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     4501 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     4561 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     4621 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     4681 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     4741 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     4801 tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa
     4861 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg
     4921 ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac
     4981 cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa
     5041 gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct
     5101 caggttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     5161 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     5221 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     5281 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     5341 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     5401 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     5461 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     5521 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     5581 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     5641 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     5701 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     5761 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     5821 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     5881 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     5941 ataaccgtat taccgccatg cat
//
