LOCUS       sequence-312944-        5865 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of self-cleaving membrane targeting peptide containing 
            the OMP25 C-terminal targeting sequence, a modified P6P4 NS5A/5B 
            cleavage site, msGFP and the NS3/4A protease..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5865)
  AUTHORS   Vevea JD, Chapman ER
  TITLE     Acute disruption of the synaptic vesicle membrane protein 
            synaptotagmin 1 using knockoff in mouse hippocampal neurons.
  JOURNAL   eLife. 2020 Jun 9;9. pii: 56469. doi: 10.7554/eLife.56469.
  PUBMED    32515733
REFERENCE   2  (bases 1 to 5865)
  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: "eLife."; 
            date: "2020-06-9"; volume: "9. pii: 56469. doi"; pages: " 
            10.7554/eLife.56469"
COMMENT     Sequence Label: pEF NS34a-msGFP-A'Q-mito
FEATURES             Location/Qualifiers
     source          1..5865
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          231..1173
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1130..1150
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1214..1223
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1934..1955)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     2542..2563
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2851..2870)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2916..2971
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2970..2989)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3332..3348)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3332..3348)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3345..3367)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3356..3372
                     /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)."
     promoter        complement(join(3380..3386,3387..3404,3405..3410))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3425..3446
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3505..3700
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3551..3686
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3706..3840
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3743..3762)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3797..3816
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3908..3925)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4079..4667)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4159..4178)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4838..5629,5630..5698))
                     /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     5461..5480
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5699..5803)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(5865,1..1182)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
ORIGIN
        1 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
       61 gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
      121 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
      181 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg
      241 tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta
      301 cttccacgcc cctggctgca gtacgtgatt cttgatcccg agcttcgggt tggaagtggg
      361 tgggagagtt cgaggccttg cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc
      421 tggcctgggc gctggggccg ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct
      481 gctttcgata agtctctagc catttaaaat ttttgatgac ctgctgcgac gctttttttc
      541 tggcaagata gtcttgtaaa tgcgggccaa gatctgcaca ctggtatttc ggtttttggg
      601 gccgcgggcg gcgacggggc ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg
      661 cgagcgcggc caccgagaat cggacggggg tagtctcaag ctggccggcc tgctctggtg
      721 cctggcctcg cgccgccgtg tatcgccccg ccctgggcgg caaggctggc ccggtcggca
      781 ccagttgcgt gagcggaaag atggccgctt cccggccctg ctgcagggag ctcaaaatgg
      841 aggacgcggc gctcgggaga gcgggcgggt gagtcaccca cacaaaggaa aagggccttt
      901 ccgtcctcag ccgtcgcttc atgtgactcc acggagtacc gggcgccgtc caggcacctc
      961 gattagttct cgagcttttg gagtacgtcg tctttaggtt ggggggaggg gttttatgcg
     1021 atggagtttc cccacactga gtgggtggag actgaagtta ggccagcttg gcacttgatg
     1081 taattctcct tggaatttgc cctttttgag tttggatctt ggttcattct caagcctcag
     1141 acagtggttc aaagtttttt tcttccattt caggtgtcgt gaggaattct gcagtcgacg
     1201 gtaccgcggg cccgccacca tgggctgcgt ggtgatcgtg ggcaggatcg tgctgagcgg
     1261 cagcggcacc agcgcgccca tcacggcgta cgcccagcag acgagaggcc tcctagggtg
     1321 tataatcacc agcctgactg gccgggacaa aaaccaagtg gagggtgagg tccagatcgt
     1381 gtcaactgct acccaaacct tcctggcaac gtgcatcaat ggggtatgct ggacagtcta
     1441 ccacggggcc ggaacgagga ccatcgcatc acccaagggt cctgtcatcc agatgtatac
     1501 caatgtggac caagaccttg tgggctggcc cgctcctcaa ggttcccgct cattgacacc
     1561 ctgtacctgc ggctcctcgg acctttacct ggtcacgagg cacgccgatg tcattcccgt
     1621 gcgccggcga ggtgatagca ggggtagcct gctttcgccc cggcccattt cctacttgaa
     1681 aggctcctcg gggggtccgc tgttgtgccc cgcgggacac gccgtgggcc tattcagggc
     1741 cgcggtgtgc acccgtggag tggctaaagc ggtggacttt atccctgtgg agaacctaga
     1801 gacaaccatg agatccccgg tgttcacgga caactcctct ccaccagcag ggtcaggttc
     1861 ttcgggcagt tcggggtcca gcggatctag tatggatagc actgagagcc tgttcaccgg
     1921 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgcg
     1981 cggcgagggc gagggcgatg ccaccaacgg caagctgacc ctgaagttca tctgcaccac
     2041 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
     2101 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     2161 aggctacgtc caggagcgca ccatcacctt caaggacgac ggcacctaca agacccgcgc
     2221 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     2281 caaggaggac ggcaacatcc tggggcacaa gctggagtac aacttcaaca gccacaacgt
     2341 ctatatcacc gccgacaagc agaagaacgg catcaaggcc aacttcaaga tccgccacaa
     2401 cgtggaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     2461 cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccaagc tgagcaaaga
     2521 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
     2581 tcacggcgga tccggaagtt cggggtccag cggttcttca ggcagttccg ctgatgtggt
     2641 ctgctgtcag atgtcctacg gcagtggatc taagcgcgga gttcctgtag ctgtggtgct
     2701 gctgccagtg tttgccctta ccctggtagc agtttgggcc ttcgtgagat accgaaagca
     2761 gctctgaggc cgcactcctc aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc
     2821 caatgccctg gctcacaaat accactgaga tctttttccc tctgccaaaa attatgggga
     2881 catcatgaag ccccttgagc atctgacttc tggctaataa aggaaattta ttttcattgc
     2941 aatagtgtgt tggaattttt tgtgtctctc actcggaagg acatatggga gggcaaatca
     3001 tttaaaacat cagaatgagt atttggttta gagtttggca acatatgccc atatgctggc
     3061 tgccatgaac aaaggttggc tataaagagg tcatcagtat atgaaacagc cccctgctgt
     3121 ccattcctta ttccatagaa aagccttgac ttgaggttag atttttttta tattttgttt
     3181 tgtgttattt ttttctttaa catccctaaa attttcctta catgttttac tagccagatt
     3241 tttcctcctc tcctgactac tcccagtcat agctgtccct cttctcttat ggagatccct
     3301 cgacctgcag cccaagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     3361 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     3421 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     3481 gaaacctgtc gtgccagcgg atccgcatct caattagtca gcaaccatag tcccgcccct
     3541 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     3601 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
     3661 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctaactt gtttattgca
     3721 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     3781 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc
     3841 cgctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     3901 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     3961 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     4021 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     4081 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4141 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     4201 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4261 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4321 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4381 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4441 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4501 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4561 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4621 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4681 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4741 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     4801 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     4861 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4921 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     4981 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     5041 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     5101 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     5161 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     5221 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     5281 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     5341 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     5401 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     5461 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5521 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     5581 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5641 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     5701 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     5761 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     5821 tgccacctgg gtcgacattg attattgact agatcatcgc gtgag
//
