LOCUS       sequence-48239-e        6128 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 6128)
  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 6128)
  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-D
FEATURES             Location/Qualifiers
     source          1..6128
                     /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      2068..2077
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2074..2076,2077..2079,2080..2793)
                     /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(2074..2076,2077..2079,2080..2790)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2119..2140)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2380..2399)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2727..2748
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2916..3037
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2953..2972)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3007..3026
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3044..3499)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3181..3202)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3393..3412
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3526..3630
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3632..3989
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3632..3823
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3655..3675)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3840..3975
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4024..4818
                     /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             4024..4818
                     /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(4078..4097)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4688..4707
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5006..5025)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5050..5097
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5426..6014
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5915..5934
                     /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 accatgacat cacgcatctt gctacgccag
      661 caactcatgc gtgagcagat gcaggagcag gagcgcaggg agcagcagca gaagctgcag
      721 gcggcccagt tcatgcaaca gagagtgccc gtgagtcaga caccagccat aaacgtcagt
      781 gtgcccacca cccttccctc tgccacgcag gtgccgatgg aagtccttaa ggtgcagacc
      841 cacctcgaaa accccaccaa gtaccacata cagcaagccc aacggcagca ggtaaagcag
      901 tacctttcta ccactttagc aaataaacat gccaaccaag tcctgagctt gccatgtcca
      961 aaccagcctg gcgatcatgt catgccaccg gtgccgggga gcagcgcacc caacagcccc
     1021 atggctatgc ttacgcttaa ctccaactgt gaaaaagagg gattttataa gtttgaagag
     1081 caaaacaggg cagagagcga gtgcccaggc atgaacacac attcacgagc gtcctgtatg
     1141 cagatggatg atgtaatcga tgacatcatt agcctagaat caagttataa tgaggaaatc
     1201 ttgggcttga tggatcctgc tttgcaaatg gcaaatacgt tgcctgtctc gggaaacttg
     1261 attgatcttt atggaaacca aggtctgccc ccaccaggcc tcaccatcag caactcctgt
     1321 ccagccaacc ttcccaacat aaaaagggag ctcacagagt ctgaagcaag agcactggcc
     1381 aaagagaggc agaaaaagga caatcacaac ctgattgaac gaagaagaag atttaacata
     1441 aatgaccgca ttaaagaact aggtactttg attcccaagt caaatgatcc agacatgcgc
     1501 tggaacaagg gaaccatctt aaaagcatcc gtggactata tccgaaagtt gcaacgagaa
     1561 cagcaacgcg caaaagaact tgaaaaccga cagaagaaac tggagcacgc caaccggcat
     1621 ttgttgctca gaatacagga acttgaaatg caggctcgag ctcatggact ttcccttatt
     1681 ccatccacgg gtctctgctc tccagatttg gtgaatcgga tcatcaagca agaacccgtt
     1741 cttgagaact gcagccaaga cctccttcag catcatgcag acctaacctg tacaacaact
     1801 ctcgatctca cggatggcac catcaccttc aacaacaacc tcggaactgg gactgaggcc
     1861 aaccaagcct atagtgtccc cacaaaaatg ggatccaaac tggaagacat cctgatggac
     1921 gacacccttt ctcccgtcgg tgtcactgat ccactccttt cctcagtgtc ccccggagct
     1981 tccaaaacaa gcagccggag gagcagtatg agcatggaag agacggagca cacttgtacg
     2041 gtaccgcggg cccgggatcc accggtcgcc accatggtga gcaagggcga ggagctgttc
     2101 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     2161 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     2221 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     2281 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     2341 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     2401 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     2461 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
     2521 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
     2581 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     2641 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
     2701 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     2761 atcactctcg gcatggacga gctgtacaag taaagcggcc gcgactctag atcataatca
     2821 gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga
     2881 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     2941 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     3001 ctagttgtgg tttgtccaaa ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat
     3061 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     3121 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     3181 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     3241 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     3301 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     3361 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     3421 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     3481 gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     3541 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     3601 atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg
     3661 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3721 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3781 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     3841 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     3901 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     3961 gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt
     4021 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     4081 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     4141 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     4201 ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     4261 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     4321 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     4381 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     4441 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     4501 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc
     4561 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     4621 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     4681 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     4741 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     4801 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     4861 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     4921 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     4981 tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc
     5041 gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata
     5101 aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg
     5161 gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc
     5221 cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata
     5281 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     5341 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     5401 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     5461 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     5521 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     5581 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     5641 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     5701 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     5761 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     5821 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     5881 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     5941 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     6001 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     6061 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     6121 ccatgcat
//
