LOCUS       sequence-449637-        5446 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of FAM177A1-GFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5446)
  AUTHORS   Ugur B, Schueder F, Shin J, Hanna MG, Wu Y, Leonzino M, Su M, McAdow
            AR, Wilson C, Postlethwait J, Solnica-Krezel L, Bewersdorf J, De 
            Camilli P
  TITLE     VPS13B is localized at the cis-trans Golgi complex interface and is 
            a functional partner of FAM177A1.
  JOURNAL   bioRxiv [Preprint]. 2023 Dec 18:2023.12.18.572081. doi: 
            10.1101/2023.12.18.572081.
  PUBMED    38187698
REFERENCE   2  (bases 1 to 5446)
  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: "bioRxiv 
            [Preprint]. 2023 Dec 18:2023.12.18.572081. doi: 
            10.1101/2023.12.18.572081."
COMMENT     Sequence Label: FAM177A1isoform2-GFP
FEATURES             Location/Qualifiers
     source          1..5446
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(175..763)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(255..274)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    1092..1139
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     1164..1183
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(1371..2165)
                     /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             complement(1371..2165)
                     /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(1482..1501)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2092..2111
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2200..2557)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2214..2349
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        2366..2557
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     2514..2534
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(2559..2663)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2690..3145
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2777..2796)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2987..3008
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    3152..3273
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3163..3182)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3217..3236
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(3396..4109,4110..4112,4113..4115))
                     /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             complement(join(3399..4109,4110..4112,4113..4115))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3441..3462)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3790..3809
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4049..4070
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      4112..4121
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      4835..4844
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(4939..5142)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4968..4988)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5143..5446
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 cggaactcca tatatgggct atgaactaat gaccccgtaa ttgattacta ttaataacta
       61 atgcatggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
      121 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
      181 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
      241 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
      301 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
      361 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
      421 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
      481 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
      541 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
      601 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
      661 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
      721 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
      781 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
      841 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
      901 ctaaagtata tatgagtaac ctgaggctat ggcagggcct gccgccccga cgttggctgc
      961 gagccctggg ccttcacccg aacttggggg gtggggtggg gaaaaggaag aaacgcgggc
     1021 gtattggccc caatggggtc tcggtggggt atcgacagag tgccagccct gggaccgaac
     1081 cccgcgttta tgaacaaacg acccaacacc gtgcgtttta ttctgtcttt ttattgccgt
     1141 catagcgcgg gttccttccg gtattgtctc cttccgtgtt tcagttagcc tccccctagg
     1201 gtgggcgaag aactccagca tgagatcccc gcgctggagg atcatccagc cggcgtcccg
     1261 gaaaacgatt ccgaagccca acctttcata gaaggcggcg gtggaatcga aatctcgtga
     1321 tggcaggttg ggcgtcgctt ggtcggtcat ttcgaacccc agagtcccgc tcagaagaac
     1381 tcgtcaagaa ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc
     1441 acgaggaagc ggtcagccca ttcgccgcca agctcttcag caatatcacg ggtagccaac
     1501 gctatgtcct gatagcggtc cgccacaccc agccggccac agtcgatgaa tccagaaaag
     1561 cggccatttt ccaccatgat attcggcaag caggcatcgc catgggtcac gacgagatcc
     1621 tcgccgtcgg gcatgctcgc cttgagcctg gcgaacagtt cggctggcgc gagcccctga
     1681 tgctcttcgt ccagatcatc ctgatcgaca agaccggctt ccatccgagt acgtgctcgc
     1741 tcgatgcgat gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag cgtatgcagc
     1801 cgccgcattg catcagccat gatggatact ttctcggcag gagcaaggtg agatgacagg
     1861 agatcctgcc ccggcacttc gcccaatagc agccagtccc ttcccgcttc agtgacaacg
     1921 tcgagcacag ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg cgctgcctcg
     1981 tcttgcagtt cattcagggc accggacagg tcggtcttga caaaaagaac cgggcgcccc
     2041 tgcgctgaca gccggaacac ggcggcatca gagcagccga ttgtctgttg tgcccagtca
     2101 tagccgaata gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc atcttgttca
     2161 atcatgcgaa acgatcctca tcctgtctct tgatcgatct ttgcaaaagc ctaggcctcc
     2221 aaaaaagcct cctcactact tctggaatag ctcagaggcc gaggcggcct cggcctctgc
     2281 ataaataaaa aaaattagtc agccatgggg cggagaatgg gcggaactgg gcggagttag
     2341 gggcgggatg ggcggagtta ggggcgggac tatggttgct gactaattga gatgcatgct
     2401 ttgcatactt ctgcctgctg gggagcctgg ggactttcca cacctggttg ctgactaatt
     2461 gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc cacaccctaa
     2521 ctgacacaca ttccacagct ggttctttcc gcctcaggac tcttcctttt tcaatattat
     2581 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     2641 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgcgccctgt
     2701 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     2761 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     2821 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     2881 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     2941 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     3001 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     3061 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     3121 aacaaaatat taacgcttac aatttacgcc ttaagataca ttgatgagtt tggacaaacc
     3181 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     3241 tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg
     3301 tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt
     3361 ggtatggctg attatgatct agagtcgcgg ccgctttact tgtacagctc gtccatgccg
     3421 agagtgatcc cggcggcggt cacgaactcc agcaggacca tgtgatcgcg cttctcgttg
     3481 gggtctttgc tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg
     3541 ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga gctgcacgct gccgtcctcg
     3601 atgttgtggc ggatcttgaa gttcaccttg atgccgttct tctgcttgtc ggccatgata
     3661 tagacgttgt ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc
     3721 ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc
     3781 tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag
     3841 ccttcgggca tggcggactt gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg
     3901 aagcactgca cgccgtaggt cagggtggtc acgagggtgg gccagggcac gggcagcttg
     3961 ccggtggtgc agatgaactt cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg
     4021 ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag gatgggcacc
     4081 accccggtga acagctcctc gcccttgctc accatggtgg cgaccggttc tggtgggaca
     4141 gagactggat tttgcttgct ttccataatt acttcactgt ctccctccat ttcaaaattg
     4201 acattcacaa atgagctgga tatcactgtc tctggttgat ctgtctgaac aatggaatca
     4261 gtctgcaatt tattctgttg atattgtttt tctgcttctt cagacatcct gttttcttct
     4321 tcttcttctt cttcctcctt cttcatccga taatattcat caatggcata ttggtacttt
     4381 ggggtgctga tacccaaaac agatgcaatc ttctctccaa ggaagtcaca cactgagaga
     4441 gttgatgtag cagcccgaag catgtaaaac cataagtagg gaccccaggt aagttttgtc
     4501 ggatcaacag taggcaaaac atctttcttc tccaggccat caacttcgtc ttcatctgtg
     4561 ctatattctt ccattgtttc accactaaca aagtggatga ctctccttgg gactttcttc
     4621 ttttttccta tgactcccag ttctacattt tcaaagcctc tttcgttact catctgccct
     4681 gcagattccc cgaatgccgc ggcggccgca gctcccgagg ctgcgacggc ttctcctcgt
     4741 tccacaccgc ccactggctc ctggtccatc gtcgtcgcca ccacagggct gctggcgctg
     4801 gtgagaaaga gggtaatggc cggtaagccc acttccatgg tggcggatcc gagctcggta
     4861 ccgtcgactg cagaattcga agcttgagct cgagatctga gtccggtagc gctagcggat
     4921 ctgacggttc actaaaccag ctctgcttat atagacctcc caccgtacac gcctaccgcc
     4981 catttgcgtc aatggggcgg agttgttacg acattttgga aagtcccgtt gattttggtg
     5041 ccaaaacaaa ctcccattga cgtcaatggg gtggagactt ggaaatcccc gtgagtcaaa
     5101 ccgctatcca cgcccattga tgtactgcca aaaccgcatc accatggtaa tagcgatgac
     5161 taatacgtag atgtactgcc aagtaggaaa gtcccataag gtcatgtact gggcataatg
     5221 ccaggcgggc catttaccgt cattgacgtc aatagggggc gtacttggca tatgatacac
     5281 ttgatgtact gccaagtggg cagtttaccg taaatactcc acccattgac gtcaatggaa
     5341 agtccctatt ggcgttacta tgggaacata cgtcattatt gacgtcaatg ggcgggggtc
     5401 gttgggcggt cagccaggcg ggccatttac cgtaagttat gtaacg
//
