LOCUS       sequence-316883-        7872 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 7872)
  AUTHORS   Venkatachalam K, Hofmann T, Montell C
  TITLE     Lysosomal localization of TRPML3 depends on TRPML2 and the 
            mucolipidosis-associated protein TRPML1.
  JOURNAL   J Biol Chem. 2006 Jun 23. 281(25):17517-27.
  PUBMED    16606612
REFERENCE   2  (bases 1 to 7872)
  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: "J Biol 
            Chem. 2006 Jun 23. 281(25):17517-27."
COMMENT     Sequence Label: TRPML1-YFP
FEATURES             Location/Qualifiers
     source          1..7872
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2459..2461,2462..3175)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2501..2522)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2762..2781)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3109..3130
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(3191..3209)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3192..3209)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3229..3246)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3235..3459
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3505..3933
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3592..3611)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3802..3823
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3942..3962)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3947..4276
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4127..4262
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4189..4208
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4343..5137
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4397..4416)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5007..5026
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5311..5432
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5348..5367)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5402..5421
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5481..5497)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5481..5497)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5494..5516)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5505..5521
                     /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(5529..5535,5536..5553,5554..5559))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5574..5595
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5712..5729)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5883..6471)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5963..5982)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6642..7433,7434..7502))
                     /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     7265..7284
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7503..7607)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7682..7701)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccgagc tcggatccac tagtccagtg tggtggaatt gcccttgcca ccatgacagc
      721 cccggcgggt ccgcgcggct cagagaccga gcggcttctg acccccaacc ccgggtatgg
      781 gacccaggcg gggccttcac cggcccctcc gacaccccca gaagaggaag accttcgccg
      841 tcgtctcaaa tactttttca tgagtccctg cgacaagttt cgagccaagg gccgcaagcc
      901 ctgcaagctg atgctgcaag tggtcaagat cctggtggtc acggtgcagc tcatcctgtt
      961 tgggctcagt aatcagctgg ctgtgacatt ccgggaagag aacaccatcg ccttccgaca
     1021 cctcttcctg ctgggctact cggacggagc ggatgacacc ttcgcagcct acacgcggga
     1081 gcagctgtac caggccatct tccatgctgt ggaccagtac ctggcgttgc ctgacgtgtc
     1141 actgggccgg tatgcgtatg tccgtggtgg gggtgaccct tggaccaatg gctcagggct
     1201 tgctctctgc cagcggtact accaccgagg ccacgtggac ccggccaacg acacatttga
     1261 cattgatccg atggtggtta ctgactgcat ccaggtggat ccccccgagc ggccccctcc
     1321 gccccccagc gacgatctca ccctcttgga aagcagctcc agttacaaga acctcacgct
     1381 caaattccac aagctggtca atgtcaccat ccacttccgg ctgaagacca ttaacctcca
     1441 gagcctcatc aataatgaga tcccggactg ctataccttc agcgtcctga tcacgtttga
     1501 caacaaagca cacagtgggc ggatccccat cagcctggag acccaggccc acatccagga
     1561 gtgtaagcac cccagtgtct tccagcacgg agacaacagc ttccggctcc tgtttgacgt
     1621 ggtggtcatc ctcacctgct ccctgtcctt cctcctctgc gcccgctcac tccttcgagg
     1681 cttcctgctg cagaacgagt ttgtggggtt catgtggcgg cagcggggac gggtcatcag
     1741 cctgtgggag cggctggaat ttgtcaatgg ctggtacatc ctgctcgtca ccagcgatgt
     1801 gctcaccatc tcgggcacca tcatgaagat cggcatcgag gccaagaact tggcgagcta
     1861 cgacgtctgc agcatcctcc tgggcacctc gacgctgctg gtgtgggtgg gcgtgatccg
     1921 ctacctgacc ttcttccaca actacaatat cctcatcgcc acactgcggg tggccctgcc
     1981 cagcgtcatg cgcttctgct gctgcgtggc tgtcatctac ctgggctact gcttctgtgg
     2041 ctggatcgtg ctggggccct atcatgtgaa gttccgctca ctctccatgg tgtctgagtg
     2101 cctgttctcg ctcatcaatg gggacgacat gtttgtgacg ttcgccgcca tgcaggcgca
     2161 gcagggccgc agcagcctgg tgtggctctt ctcccagctc tacctttact ccttcatcag
     2221 cctcttcatc tacatggtgc tcagcctctt catcgcgctc atcaccggcg cctacgacac
     2281 catcaagcat cccggcggcg caggcgcaga ggagagcgag ctgcaggcct acatcgcaca
     2341 gtgccaggac agccccacct ccggcaagtt ccgccgcggg agcggctcgg cctgcagcct
     2401 tctctgctgc tgcggaaggg acccctcgga ggagcattcg ctgctggtga atctcgaggt
     2461 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     2521 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     2581 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     2641 gaccaccttc ggctacggcc tgcagtgctt cgcccgctac cccgaccaca tgaagcagca
     2701 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     2761 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     2821 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     2881 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     2941 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     3001 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     3061 gagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     3121 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaatctag
     3181 agggccctat tctatagtgt cacctaaatg ctagagctcg ctgatcagcc tcgactgtgc
     3241 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     3301 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
     3361 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     3421 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     3481 gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     3541 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     3601 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     3661 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     3721 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     3781 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     3841 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     3901 atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg
     3961 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     4021 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     4081 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     4141 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     4201 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     4261 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga
     4321 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
     4381 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     4441 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     4501 gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac
     4561 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     4621 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     4681 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     4741 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     4801 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     4861 gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     4921 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     4981 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     5041 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     5101 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     5161 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     5221 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     5281 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
     5341 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     5401 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
     5461 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     5521 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     5581 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     5641 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     5701 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     5761 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     5821 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     5881 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     5941 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     6001 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     6061 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     6121 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     6181 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     6241 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     6301 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     6361 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     6421 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     6481 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     6541 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     6601 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     6661 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     6721 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     6781 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     6841 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     6901 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     6961 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     7021 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     7081 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     7141 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     7201 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     7261 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     7321 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     7381 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     7441 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     7501 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     7561 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     7621 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
     7681 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
     7741 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
     7801 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
     7861 gatatacgcg tt
//
