LOCUS       sequence-322909-        5650 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of human TMEM192. Dileucine motifs LL23-24 and 
            LL28-29 have been mutated to AA..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5650)
  AUTHORS   Adelmann CH, Traunbauer AK, Chen B, Condon KJ, Chan SH, Kunchok T, 
            Lewis CA, Sabatini DM
  TITLE     MFSD12 mediates the import of cysteine into melanosomes and 
            lysosomes.
  JOURNAL   Nature. 2020 Nov 18. pii: 10.1038/s41586-020-2937-x. doi: 
            10.1038/s41586-020-2937-x.
  PUBMED    33208952
REFERENCE   2  (bases 1 to 5650)
  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: "Nature. 
            2020 Nov 18. pii: 10.1038/s41586-020-2937-x. doi: 
            10.1038/s41586-020-2937-x."
COMMENT     Sequence Label: prk5-TMEM192-3xHA PM mutant
FEATURES             Location/Qualifiers
     source          1..5650
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      931..940
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      935..944
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1765..1791
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1804..1830
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1843..1869
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1906..1929
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      1950..1959
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1963..2097
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2000..2019)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2054..2073
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2166..2523
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2166..2357
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2189..2209)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2374..2509
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2543..2560)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2543..2559)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2552..2574)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2772..3227
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2859..2878)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3069..3090
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3244..3263)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3363..3385
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3423..3441)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3509..3613
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3614..3682,3683..4474)
                     /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(3832..3851)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4645..5233
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5134..5153
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5387..5404
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5521..5542
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5557..5562,5563..5580,5581..5587)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5595..5611
                     /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     5600..5622
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5619..5635
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5619..5635
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attcgtcgac gccaccatgg cggcgggggg caggatggag
      961 gacggttcct tggatatcac ccagagtatt gaagacgacc cagcggctga tgcccaggct
     1021 gctccacacc actcattaca agctcacttt agaccccgat tccatcctct tcctacagtc
     1081 atcatagtga atcttctgtg gtttattcat ctcgtgtttg ttgttttagc atttttaaca
     1141 ggtgtgcttt gttcttatcc taatccaaat gaggacaagt gcccaggaaa ttacacaaac
     1201 ccattgaaag ttcagacggt tataatcctt gggaaagtta ttttgtggat tctccattta
     1261 ctccttgaat gctacatcca gtatcaccac agcaaaatca gaaaccgagg ctataacttg
     1321 atctaccgat caacaaggca tctcaagaga cttgcgttga tgatacagtc ctctggcaac
     1381 acagtgcttc tcctcatact gtgcatgcag cactccttcc cagagcctgg cagattgtat
     1441 cttgacctca ttctggccat cttggcactg gaactcatct gttccctgat atgtctcctc
     1501 atttacacag tgaaaatccg gagatttaat aaagctaaac cagagcctga tatacttgaa
     1561 gaagaaaaaa tctatgctta ccccagcaat attacctcgg agactggatt cagaactatt
     1621 tcaagcctag aagaaattgt tgaaaagcaa ggagacacca ttgaatacct gaagcgacac
     1681 aatgcgctgc tgagtaagcg attgttggct ctcacttcct cagacctggg ctgtcagcca
     1741 agtagaacgg gaggtggagg tagttatccc tatgacgtgc ctgattacgc cggcacagga
     1801 tcctacccct atgatgtgcc tgactacgct ggcagcgccg gataccctta tgatgtgcct
     1861 gattatgctg gagggagcgg ctaagcggcc gcaggaggtg gaggtgacta caaggatgac
     1921 gatgacaagt gataaggatc cccagcttgg ccgccatggc ccaacttgtt tattgcagct
     1981 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     2041 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatcggg
     2101 aattaattcg gcgcagcacc atggcctgaa ataacctctg aaagaggaac ttggttaggt
     2161 accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     2221 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     2281 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2341 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     2401 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     2461 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     2521 aaaaagctgt taacagcttg gcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     2581 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     2641 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc
     2701 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa
     2761 gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2821 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2881 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2941 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc
     3001 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     3061 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc
     3121 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     3181 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct
     3241 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc
     3301 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     3361 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa
     3421 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac
     3481 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     3541 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     3601 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     3661 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     3721 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     3781 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     3841 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     3901 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     3961 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     4021 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     4081 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     4141 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     4201 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     4261 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     4321 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     4381 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     4441 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     4501 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     4561 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     4621 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     4681 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     4741 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     4801 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     4861 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     4921 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     4981 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     5041 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     5101 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     5161 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     5221 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     5281 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     5341 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     5401 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     5461 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     5521 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
     5581 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgacatgat
     5641 tacgaattaa
//
