LOCUS       sequence-190817-        5635 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of mutant TMEM230 (isoform 2) and ZsGreen.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5635)
  AUTHORS   Deng HX, Shi Y, Yang Y, Ahmeti KB, Miller N, Huang C, Cheng L, Zhai 
            H, Deng S, Nuytemans K, Corbett NJ, Kim MJ, Deng H, Tang B, Yang Z, 
            Xu Y, Chan P, Huang B, Gao XP, Song Z, Liu Z, Fecto F, Siddique N, 
            Foroud T, Jankovic J, Ghetti B, Nicholson DA, Krainc D, Melen O, 
            Vance JM, Pericak-Vance MA, Ma YC, Rajput AH, Siddique T
  TITLE     Identification of TMEM230 mutations in familial Parkinson's disease.
  JOURNAL   Nat Genet. 2016 Jul;48(7):733-9. doi: 10.1038/ng.3589. Epub 2016 Jun
            6.
  PUBMED    27270108
REFERENCE   2  (bases 1 to 5635)
  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: "10.1038/ng.3589"; 
            journalName: "Nat Genet"; date: "2016-07"; volume: "48"; issue: "7";
            pages: "733-9"
COMMENT     Sequence Label: TMEM230-M4/X184PG-IRES(isoform 2)
FEATURES             Location/Qualifiers
     source          1..5635
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      25..613
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     514..533
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        788..1091
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1092..1295
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1246..1266
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    join(1746..2320,2321..2323,2324..2332)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1913..1930)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2140..2159
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2333..3028
                     /codon_start=1
                     /product="Zoanthus green fluorescent protein"
                     /label=ZsGreen1
                     /note="mammalian codon-optimized"
                     /translation="MAQSKHGLTKEMTMKYRMEGCVDGHKFVITGEGIGYPFKGKQAIN
                     LCVVEGGPLPFAEDILSAAFMYGNRVFTEYPQDIVDYFKNSCPAGYTWDRSFLFEDGAV
                     CICNADITVSVEENCMYHESKFYGVNFPADGPVMKKMTDNWEPSCEKIIPVPKQGILKG
                     DVSMYLLLKDGGRLRCQFDTVYKAKSVPRKMPDWHFIQHKLTREDRSDAKNQKWHLTEH
                     AIASGSALP"
     polyA_signal    3150..3271
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3187..3206)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3241..3260
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3278..3733)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3415..3436)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3627..3646
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3760..3864
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3866..4223
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3866..4057
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3889..3909)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4074..4209
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4258..5052
                     /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             4258..5052
                     /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(4312..4331)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4922..4941
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5240..5259)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5284..5331
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
       61 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
      121 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
      181 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
      241 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
      301 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
      361 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
      421 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
      481 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
      541 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
      601 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
      661 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
      721 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
      781 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      841 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      901 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      961 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
     1021 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
     1081 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
     1141 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
     1201 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
     1261 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
     1321 agcgctaccg gactcagatc tcgaggccac catgatgccg tcccgtacca acctggctac
     1381 tggaatcccc agtagtaaag tgaaatattc aaggctctcc agcacagacg atggctacat
     1441 tgaccttcag tttaagaaaa cccctcctaa gatcccttat aaggccatcg cacttgccac
     1501 tgtgctgttt ttgattggcg cctttctcat tattataggc tccctcctgc tgtcaggcta
     1561 catcagcaaa gggggggcag accgggccgt tccagtgctg atcattggca ttctggtgtt
     1621 cctacccgga ttttaccacc tgcgcatcgc ttactatgca tccaaaggct accgtggtta
     1681 ctcctatgat gacattccag actttgatga ccccgggcac ccaccccata gctgaggagg
     1741 atccgcccct ctccctcccc cccccctaac gttactggcc gaagccgctt ggaataaggc
     1801 cggtgtgcgt ttgtctatat gttattttcc accatattgc cgtcttttgg caatgtgagg
     1861 gcccggaaac ctggccctgt cttcttgacg agcattccta ggggtctttc ccctctcgcc
     1921 aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag ttcctctgga agcttcttga
     1981 agacaaacaa cgtctgtagc gaccctttgc aggcagcgga accccccacc tggcgacagg
     2041 tgcctctgcg gccaaaagcc acgtgtataa gatacacctg caaaggcggc acaaccccag
     2101 tgccacgttg tgagttggat agttgtggaa agagtcaaat ggctctcctc aagcgtattc
     2161 aacaaggggc tgaaggatgc ccagaaggta ccccattgta tgggatctga tctggggcct
     2221 cggtacacat gctttacatg tgtttagtcg aggttaaaaa aacgtctagg ccccccgaac
     2281 cacggggacg tggttttcct ttgaaaaaca cgatgataat atggccacaa ccatggccca
     2341 gtccaagcac ggcctgacca aggagatgac catgaagtac cgcatggagg gctgcgtgga
     2401 cggccacaag ttcgtgatca ccggcgaggg catcggctac cccttcaagg gcaagcaggc
     2461 catcaacctg tgcgtggtgg agggcggccc cttgcccttc gccgaggaca tcttgtccgc
     2521 cgccttcatg tacggcaacc gcgtgttcac cgagtacccc caggacatcg tcgactactt
     2581 caagaactcc tgccccgccg gctacacctg ggaccgctcc ttcctgttcg aggacggcgc
     2641 cgtgtgcatc tgcaacgccg acatcaccgt gagcgtggag gagaactgca tgtaccacga
     2701 gtccaagttc tacggcgtga acttccccgc cgacggcccc gtgatgaaga agatgaccga
     2761 caactgggag ccctcctgcg agaagatcat ccccgtgccc aagcagggca tcttgaaggg
     2821 cgacgtgagc atgtacctgc tgctgaagga cggtggccgc ttgcgctgcc agttcgacac
     2881 cgtgtacaag gccaagtccg tgccccgcaa gatgcccgac tggcacttca tccagcacaa
     2941 gctgacccgc gaggaccgca gcgacgccaa gaaccagaag tggcacctga ccgagcacgc
     3001 catcgcctcc ggctccgcct tgccctgagc ggccgcgact ctagatcata atcagccata
     3061 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     3121 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     3181 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     3241 gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg
     3301 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     3361 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     3421 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     3481 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     3541 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     3601 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     3661 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     3721 ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     3781 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     3841 caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc
     3901 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3961 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     4021 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     4081 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4141 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     4201 ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg
     4261 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     4321 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4381 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa
     4441 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4501 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4561 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     4621 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4681 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4741 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc
     4801 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4861 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4921 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4981 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     5041 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     5101 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     5161 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     5221 accctagggg gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta
     5281 tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg
     5341 gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat
     5401 acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc
     5461 tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt
     5521 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     5581 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccc
//
