LOCUS       sequence-449636-        7067 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of ZFPL1-GFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7067)
  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 7067)
  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: ZFPL1-GFP
FEATURES             Location/Qualifiers
     source          1..7067
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(65..859)
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(176..195)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     786..805
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(926..1255)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      940..1075
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(994..1013)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     1240..1260
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(1269..1697)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1379..1400)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1591..1610
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    1743..1967
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     1956..1973
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(join(2005..2718,2719..2721))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             complement(2008..2721)
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(2050..2071)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2399..2418
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2658..2679
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3705..3714
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3749..3768)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3750..3768)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(3813..4016)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3842..3862)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4017..4396
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     4568..4587
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4662..4766
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4767..4835,4836..5627)
                     /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(4985..5004)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5798..6386
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6287..6306
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6540..6557
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6674..6695
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6710..6715,6716..6733,6734..6740)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6748..6764
                     /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     6753..6775
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6772..6788
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6772..6788
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    6837..6958
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6848..6867)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6902..6921
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
ORIGIN
        1 tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg tcatttcgaa ccccagagtc
       61 ccgctcagaa gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa tcgggagcgg
      121 cgataccgta aagcacgagg aagcggtcag cccattcgcc gccaagctct tcagcaatat
      181 cacgggtagc caacgctatg tcctgatagc ggtccgccac acccagccgg ccacagtcga
      241 tgaatccaga aaagcggcca ttttccacca tgatattcgg caagcaggca tcgccatggg
      301 tcacgacgag atcctcgccg tcgggcatgc gcgccttgag cctggcgaac agttcggctg
      361 gcgcgagccc ctgatgctct tcgtccagat catcctgatc gacaagaccg gcttccatcc
      421 gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc gaatgggcag gtagccggat
      481 caagcgtatg cagccgccgc attgcatcag ccatgatgga tactttctcg gcaggagcaa
      541 ggtgagatga caggagatcc tgccccggca cttcgcccaa tagcagccag tcccttcccg
      601 cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc cgtcgtggcc agccacgata
      661 gccgcgctgc ctcgtcctgc agttcattca gggcaccgga caggtcggtc ttgacaaaaa
      721 gaaccgggcg cccctgcgct gacagccgga acacggcggc atcagagcag ccgattgtct
      781 gttgtgccca gtcatagccg aatagcctct ccacccaagc ggccggagaa cctgcgtgca
      841 atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt ctcttgatca gatccgaaaa
      901 tggatataca agctcccggg agctttttgc aaaagcctag gcctccaaaa aagcctcctc
      961 actacttctg gaatagctca gaggcagagg cggcctcggc ctctgcataa ataaaaaaaa
     1021 ttagtcagcc atggggcgga gaatgggcgg aactgggcgg agttaggggc gggatgggcg
     1081 gagttagggg cgggactatg gttgctgact aattgagatg catgctttgc atacttctgc
     1141 ctgctgggga gcctggggac tttccacacc tggttgctga ctaattgaga tgcatgcttt
     1201 gcatacttct gcctgctggg gagcctgggg actttccaca ccctaactga cacacattcc
     1261 acagaattaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
     1321 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
     1381 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
     1441 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
     1501 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
     1561 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
     1621 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
     1681 gccgctacag ggcgcgtggg gataccccct agagccccag ctggttcttt ccgcctcaga
     1741 agccatagag cccaccgcat ccccagcatg cctgctattg tcttcccaat cctccccctt
     1801 gctgtcctgc cccaccccac cccccagaat agaatgacac ctactcagac aatgcgatgc
     1861 aatttcctca ttttattagg aaaggacagt gggagtggca ccttccaggg tcaaggaagg
     1921 cacgggggag gggcaaacaa cagatggctg gcaactagaa ggcacagtcg aggctgatca
     1981 gcgggtttaa acgggccctc tagattactt gtacagctcg tccatgccgt gagtgatccc
     2041 ggcggcggtc acgaactcca gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct
     2101 cagggcggac tgggtgctca ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat
     2161 gggggtgttc tgctggtagt ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg
     2221 gatcttgaag ttcaccttga tgccgttctt ctgcttgtcg gccatgatat agacgttgtg
     2281 gctgttgtag ttgtactcca gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat
     2341 gcccttcagc tcgatgcggt tcaccagggt gtcgccctcg aacttcacct cggcgcgggt
     2401 cttgtagttg ccgtcgtcct tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat
     2461 ggcggacttg aagaagtcgt gctgcttcat gtggtcgggg tagcggctga agcactgcac
     2521 gccgtaggtc agggtggtca cgagggtggg ccagggcacg ggcagcttgc cggtggtgca
     2581 gatgaacttc agggtcagct tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct
     2641 gaacttgtgg ccgtttacgt cgccgtccag ctcgaccagg atgggcacca ccccggtgaa
     2701 cagctcctcg cccttgctca tgcggccgcc actgtgctgg atatctgcag aattccacca
     2761 cactggacta gtggatccgg aggggcccac gcggatgtga gggttcatga gtgggtccag
     2821 gttgggatcg ctgtcagctg cggcccggcc taggcgagac atgagggcaa ggagggccag
     2881 gaagcccagc agtcccaaga gtagcagcag ccccgcccgc tggagcaggg tcagcggccg
     2941 cttccgagac ccagcccggc tccttagcag ccgggccagc caacccaagg ccggccgacg
     3001 tcggtacttg tcatcgtcac agtctccatg gaggcctggt gtccggtcat catcccgcgt
     3061 atcatacacc ttcctagggg cgtgagtcaa gggctcagga ttgcccatgt ggatcactgt
     3121 gtgctgctcg ggccggcctg gggaagctgg gggccggggg gcctggctgt agaaggctgg
     3181 ggcagcagag gcgctgtcta cctcctctgg tccaggggta ctgctggcat taaaactaga
     3241 ccagtcagag aagtcagacg tgttgagggg ctcgggctct gggctcacca cctcatcgat
     3301 cagagggagg cccagtcctg cccgggccca gttgactgtg gccagcttct ctctcagtgc
     3361 ggaggccacg gggccagcca ggttggttgg ggggaagatg gggccattgc agctggggca
     3421 ctgatagccg gcaggtgccg tgtttcgggg tagctgggca gcacgttcat tgaggcaggc
     3481 ccagtgaaag agatcatagc agacaaggcg ggtcgtctct cggctggcca ggggtatgtt
     3541 gcacaggcgg caattggggt tgtagtcgct atcttggagc cattgcaggt aggactggac
     3601 gatgcacttg gcgtgattgg ctaccaggca gtgctcgcag acgttgaccc gatgttcgaa
     3661 gcagaacagg ttggtcacct ttctcttggg gcacttacaa agccccatgg tggcaagctt
     3721 aagtttaaac gctagccagc ttgggtctcc ctatagtgag tcgtattaat ttcgataagc
     3781 cagtaagcag tgggttctct agttagccag agagctctgc ttatatagac ctcccaccgt
     3841 acacgcctac cgcccatttg cgtcaatggg gcggagttgt tacgacattt tggaaagtcc
     3901 cgttgatttt ggtgccaaaa caaactccca ttgacgtcaa tggggtggag acttggaaat
     3961 ccccgtgagt caaaccgcta tccacgccca ttgatgtact gccaaaaccg catcaccatg
     4021 gtaatagcga tgactaatac gtagatgtac tgccaagtag gaaagtccca taaggtcatg
     4081 tactgggcat aatgccaggc gggccattta ccgtcattga cgtcaatagg gggcgtactt
     4141 ggcatatgat acacttgatg tactgccaag tgggcagttt accgtaaata ctccacccat
     4201 tgacgtcaat ggaaagtccc tattggcgtt actatgggaa catacgtcat tattgacgtc
     4261 aatgggcggg ggtcgttggg cggtcagcca ggcgggccat ttaccgtaag ttatgtaacg
     4321 cggaactcca tatatgggct atgaactaat gaccccgtaa ttgattacta ttaataacta
     4381 gtcaataatc aatgtcaacg cgtatatctg gcccgtacat cgcgaagcag cgcaaaacgc
     4441 ctaaccctaa gcagattctt catgcaattg tcggtcaagc cttgccttgt tgtagcttaa
     4501 attttgctcg cgcactactc agcgacctcc aacacacaag cagggagcag atactggctt
     4561 aactatgcgg catcagagca gattgtactg agagtgcacc ataggggatc gggagatctc
     4621 ccgatccgtc gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     4681 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     4741 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     4801 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     4861 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     4921 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     4981 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     5041 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     5101 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     5161 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     5221 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     5281 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     5341 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     5401 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     5461 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     5521 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     5581 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     5641 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     5701 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     5761 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     5821 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     5881 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     5941 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     6001 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     6061 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     6121 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     6181 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     6241 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     6301 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     6361 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     6421 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     6481 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     6541 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     6601 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     6661 agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta
     6721 tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca
     6781 gctatgacca tgattacgcc aagctctagc tagaggtcga cggtatacag acatgataag
     6841 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     6901 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttgg
     6961 ggtgggcgaa gaactccagc atgagatccc cgcgctggag gatcatccag ccggcgtccc
     7021 ggaaaacgat tccgaagccc aacctttcat agaaggcggc ggtggaa
//
