LOCUS       sequence-443321-        6260 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TMEM9 expression vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6260)
  AUTHORS   Gonzalez-Lozano MA, Schmid EW, Miguel Whelan E, Jiang Y, Paulo JA, 
            Walter JC, Harper JW
  TITLE     EndoMAP.v1 charts the structural landscape of human early endosome 
            complexes.
  JOURNAL   Nature. 2025 Jul;643(8070):252-261. doi: 10.1038/s41586-025-09059-y.
            Epub 2025 May 28.
  PUBMED    40437099
REFERENCE   2  (bases 1 to 6260)
  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/s41586-025-09059-y"; journalName: "Nature"; date: 
            "2025-07"; volume: "643"; issue: "8070"; pages: "252-261"
COMMENT     Sequence Label: pcDNA -TMEM9-3xHA
FEATURES             Location/Qualifiers
     source          1..6260
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1040..1161
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1051..1070)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1105..1124
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1291..2316)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(1291..2316)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     promoter        complement(2365..2694)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2379..2514
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2433..2452)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     2679..2699
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(2708..3136)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2818..2839)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3030..3049
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    3285..3396
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3385..3402
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(3445..3525)
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     protein_bind    3527..3551
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(4101..4125)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(4212..4231)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4213..4231)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(4276..4479)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4305..4325)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4480..4859
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5031..5050
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5125..5229
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5230..5298,5299..6090)
                     /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(5448..5467)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct agctagaggt
     1021 cgacggtata cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     1081 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     1141 ataagctgca ataaacaagt tggggtgggc gaagaactcc agcatgagat ccccgcgctg
     1201 gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc
     1261 ggcggtggaa tcgaaatctc gtagcacgtg ctattccttt gccctcggac gagtgctggg
     1321 gcgtcggttt ccactatcgg cgagtacttc tacacagcca tcggtccaga cggccgcgct
     1381 tctgcgggcg atttgtgtac gcccgacagt cccggctccg gatcggacga ttgcgtcgca
     1441 tcgaccctgc gcccaagctg catcatcgaa attgccgtca accaagctct gatagagttg
     1501 gtcaagacca atgcggagca tatacgcccg gagccgcggc gatcctgcaa gctccggatg
     1561 cctccgctcg aagtagcgcg tctgctgctc catacaagcc aaccacggcc tccagaagaa
     1621 gatgttggcg acctcgtatt gggaatcccc gaacatcgcc tcgctccagt caatgaccgc
     1681 tgttatgcgg ccattgtccg tcaggacatt gttggagccg aaatccgcgt gcacgaggtg
     1741 ccggacttcg gggcagtcct cggcccaaag catcagctca tcgagagcct gcgcgacgga
     1801 cgcactgacg gtgtcgtcca tcacagtttg ccagtgatac acatggggat cagcaatcgc
     1861 gcatatgaaa tcacgccatg tagtgtattg accgattcct tgcggtccga atgggccgaa
     1921 cccgctcgtc tggctaagat cggccgcagc gatcgcatcc atggcctccg cgaccggctg
     1981 cagaacagcg ggcagttcgg tttcaggcag gtcttgcaac gtgacaccct gtgcacggcg
     2041 ggagatgcaa taggtcaggc tctcgctgaa ttccccaatg tcaagcactt ccggaatcgg
     2101 gagcgcggcc gatgcaaagt gccgataaac ataacgatct ttgtagaaac catcggcgca
     2161 gctatttacc cgcaggacat atccacgccc tcctacatcg aagctgaaag cacgagattc
     2221 ttcgccctcc gagagctgca tcaggtcgga gacgctgtcg aacttttcga tcagaaactt
     2281 ctcgacagac gtcgcggtga gttcaggctt tttcatcacg tgctgatcag atccgaaaat
     2341 ggatatacaa gctcccggga gctttttgca aaagcctagg cctccaaaaa agcctcctca
     2401 ctacttctgg aatagctcag aggcagaggc ggcctcggcc tctgcataaa taaaaaaaat
     2461 tagtcagcca tggggcggag aatgggcgga actgggcgga gttaggggcg ggatgggcgg
     2521 agttaggggc gggactatgg ttgctgacta attgagatgc atgctttgca tacttctgcc
     2581 tgctggggag cctggggact ttccacacct ggttgctgac taattgagat gcatgctttg
     2641 catacttctg cctgctgggg agcctgggga ctttccacac cctaactgac acacattcca
     2701 cagaattaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     2761 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     2821 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     2881 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     2941 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     3001 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     3061 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     3121 ccgctacagg gcgcgtgggg atacccccta gagccccagc tggttctttc cgcctcagaa
     3181 gccatagagc ccaccgcatc cccagcatgc ctgctattgt cttccccttg ctgtcctgcc
     3241 ccaccccacc ccccagaata gaatgacacc tactcagaca atgcgatgca atttcctcat
     3301 tttattagga aaggacagtg ggagtggcac cttccagggt caaggaaggc acgggggagg
     3361 ggcaaacaac agatggctgg caactagaag gcacagtcga ggctgatcag cgggtttaaa
     3421 cgggccctct agactcgagt ttcaagcgta gtctggtaca tcataagggt atgcatagtc
     3481 aggtacgtca taaggataag cgtaatctgg aacatcgtat gggtaaacca ctttgtacaa
     3541 gaaagttggg tagctgagca tcttgtgccg atcgaagact gtcttccgct gctcctgcac
     3601 ctgcagcttc caccgctgct gggcaccttc cacacgctcc aggactgtgt ttgctcgggg
     3661 tcccccgagg gatgcagcag ctgctgccat agagcgagca tcctcattct cctcctcatt
     3721 gtgcagttgc tcagtgtatg catccggctt tcggatcaga gggtccacca gcatcaggaa
     3781 ggccatgtag agcaacaggg cacccaccac ggacaggtag atgacaatga tgaccttgat
     3841 ggtggtggtg ctgcgctcct cgtacctgca ctcgcacagc aggcagtagg cctccacgtc
     3901 atggccaggc actggcatgg gctccaccac gtgcaggcag ttgcagtcct tctgggatac
     3961 attctggttg taaatgtgcc cactgatgtt tctataaggt ggacagatgc atttgcaccg
     4021 gatatcttca gaactcttgt tggcttcagc tgggggcacc agcaaacacc cgaccacagc
     4081 caccaaagat aagagcttca tgccaacttt tttgtacaaa cttgttgata tctgcagaat
     4141 tccaccacac tggactagtg gatccgagct cggtaccaag cttaagttta aacgctagcc
     4201 agcttgggtc tccctatagt gagtcgtatt aatttcgata agccagtaag cagtgggttc
     4261 tctagttagc cagagagctc tgcttatata gacctcccac cgtacacgcc taccgcccat
     4321 ttgcgtcaat ggggcggagt tgttacgaca ttttggaaag tcccgttgat tttggtgcca
     4381 aaacaaactc ccattgacgt caatggggtg gagacttgga aatccccgtg agtcaaaccg
     4441 ctatccacgc ccattgatgt actgccaaaa ccgcatcacc atggtaatag cgatgactaa
     4501 tacgtagatg tactgccaag taggaaagtc ccataaggtc atgtactggg cataatgcca
     4561 ggcgggccat ttaccgtcat tgacgtcaat agggggcgta cttggcatat gatacacttg
     4621 atgtactgcc aagtgggcag tttaccgtaa atactccacc cattgacgtc aatggaaagt
     4681 ccctattggc gttactatgg gaacatacgt cattattgac gtcaatgggc gggggtcgtt
     4741 gggcggtcag ccaggcgggc catttaccgt aagttatgta acgcggaact ccatatatgg
     4801 gctatgaact aatgaccccg taattgatta ctattaataa ctagtcaata atcaatgtca
     4861 acgcgtatat ctggcccgta catcgcgaag cagcgcaaaa cgcctaaccc taagcagatt
     4921 cttcatgcaa ttgtcggtca agccttgcct tgttgtagct taaattttgc tcgcgcacta
     4981 ctcagcgacc tccaacacac aagcagggag cagatactgg cttaactatg cggcatcaga
     5041 gcagattgta ctgagagtgc accatagggg atcgggagat ctcccgatcc gtcgacgtca
     5101 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     5161 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     5221 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5281 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5341 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5401 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     5461 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5521 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5581 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5641 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     5701 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     5761 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     5821 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     5881 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     5941 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     6001 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     6061 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     6121 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6181 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6241 gaaaagatca aaggatcttc
//
