LOCUS       sequence-256542-        6400 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human TMEM206 tagged at the C-terminus with FLAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6400)
  AUTHORS   Yang J, Chen J, Del Carmen Vitery M, Osei-Owusu J, Chu J, Yu H, Sun 
            S, Qiu Z
  TITLE     PAC, an evolutionarily conserved membrane protein, is a 
            proton-activated chloride channel.
  JOURNAL   Science. 2019 Apr 26;364(6438):395-399. doi: 
            10.1126/science.aav9739. Epub 2019 Apr 25.
  PUBMED    31023925
REFERENCE   2  (bases 1 to 6400)
  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"; doi: 
            "10.1126/science.aav9739"; journalName: "Science"; date: 
            "2019-04-26- 26"; volume: "364"; issue: "6438"; pages: "395-399"
COMMENT     Sequence Label: pIRES2-hPAC-FLAG
FEATURES             Location/Qualifiers
     source          1..6400
                     /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"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1671..1694
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    join(1759..2333,2334..2336,2337..2345)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1926..1943)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2153..2172
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2346..2348,2349..2351,2352..3065)
                     /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(2346..2348,2349..2351,2352..3062)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2391..2412)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2652..2671)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2999..3020
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3188..3309
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3225..3244)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3279..3298
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3316..3771)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3453..3474)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3665..3684
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3798..3902
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3904..4261
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3904..4095
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3927..3947)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4112..4247
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4296..5090
                     /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             4296..5090
                     /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(4350..4369)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4960..4979
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5278..5297)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5322..5369
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5698..6286
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6187..6206
                     /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 atctcgagcc atgatccggc aggagcgctc cacatcctac caggagctga
      661 gtgaggagtt ggtccaggtg gttgagaact cagagctggc agacgagcag gacaaggaga
      721 cggtcagagt ccaaggtccg ggtatcttac caggcctgga cagcgagtcc gcctccagca
      781 gcatccgctt cagcaaggcc tgcctgaaga acgtcttctc ggtcctactc atcttcatct
      841 acctgctgct catggctgtg gccgtcttcc tggtctaccg gaccatcaca gactttcgtg
      901 agaaactcaa gcaccctgtc atgtctgtgt cttacaagga agtggatcgc tatgatgccc
      961 caggtattgc cttgtacccc ggtcaggccc agttgctcag ctgtaagcac cattacgagg
     1021 tcattcctcc tctgacaagc cctggccagc cgggtgacat gaattgcacc acccagagga
     1081 tcaactacac ggaccccttc tccaatcaga ctgtgaaatc tgccctgatt gtccaggggc
     1141 cccgggaagt gaaaaagcgg gagctggtct tcctccagtt ccgcctgaac aagagtagtg
     1201 aggacttcag cgccattgat tacctcctct tctcttcttt ccaggagttc ctgcaaagcc
     1261 caaacagggt aggcttcatg caggcctgtg agagtgccta ttccagctgg aagttctctg
     1321 ggggcttccg cacctgggtc aagatgtcac tggtaaagac caaggaggag gatgggcggg
     1381 aagcagtgga gttccggcag gagacaagtg tggttaacta cattgaccag aggccagctg
     1441 ccaaaaaaag tgctcaattg ttttttgtgg tctttgaatg gaaagatcct ttcatccaga
     1501 aagtccaaga tatagtcact gccaatcctt ggaacacaat tgctcttctc tgtggcgcct
     1561 tcttggcatt atttaaagca gcagagtttg ccaaactgag tataaaatgg atgatcaaaa
     1621 ttagaaagag ataccttaaa agaagaggtc aggcaacgag ccacataagc gactacaaag
     1681 acgatgacga caagtgagaa ttctgcagtc gacaccggtg cgatcgcacg cgtccgcggc
     1741 ccgggcgcgc cggatccgcc cctctccctc ccccccccct aacgttactg gccgaagccg
     1801 cttggaataa ggccggtgtg cgtttgtcta tatgttattt tccaccatat tgccgtcttt
     1861 tggcaatgtg agggcccgga aacctggccc tgtcttcttg acgagcattc ctaggggtct
     1921 ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct
     1981 ggaagcttct tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc ggaacccccc
     2041 acctggcgac aggtgcctct gcggccaaaa gccacgtgta taagatacac ctgcaaaggc
     2101 ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg gaaagagtca aatggctctc
     2161 ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag gtaccccatt gtatgggatc
     2221 tgatctgggg cctcggtaca catgctttac atgtgtttag tcgaggttaa aaaaacgtct
     2281 aggccccccg aaccacgggg acgtggtttt cctttgaaaa acacgatgat aatatggcca
     2341 caaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2401 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2461 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2521 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     2581 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     2641 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     2701 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     2761 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     2821 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     2881 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     2941 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     3001 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3061 agtaaagcgg ccgcgactct agatcataat cagccatacc acatttgtag aggttttact
     3121 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     3181 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     3241 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     3301 tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     3361 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     3421 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     3481 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     3541 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     3601 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     3661 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     3721 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     3781 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     3841 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     3901 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     3961 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     4021 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4081 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     4141 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     4201 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     4261 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     4321 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     4381 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     4441 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     4501 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     4561 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     4621 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     4681 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     4741 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     4801 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     4861 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     4921 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     4981 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     5041 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     5101 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     5161 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     5221 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     5281 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     5341 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     5401 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     5461 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     5521 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     5581 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     5641 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5701 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5761 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5821 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     5881 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5941 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     6001 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6061 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     6121 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     6181 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     6241 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     6301 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     6361 atcccctgat tctgtggata accgtattac cgccatgcat
//
