LOCUS       sequence-355802-        7317 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Mammalian expression of VAMP2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7317)
  AUTHORS   Nabb AT, Bentley M
  TITLE     NgCAM and VAMP2 Reveal that Direct Delivery and Dendritic 
            Degradation Maintain Axonal Polarity.
  JOURNAL   Mol Biol Cell. 2021 Nov 3:mbcE21080425. doi: 
            10.1091/mbc.E21-08-0425.
  PUBMED    34731031
REFERENCE   2  (bases 1 to 7317)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Biol 
            Cell. 2021 Nov 3:mbcE21080425. doi: 10.1091/mbc.E21-08-0425."
COMMENT     Sequence Label: pBa.VAMP2-GFP-SBP
FEATURES             Location/Qualifiers
     source          1..7317
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..276
                     /label=chicken beta-actin promoter
     primer_bind     1219..1241
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     regulatory      1705..1714
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1711..1713,1714..1716,1717..2424)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY"
     CDS             join(1711..1713,1714..1716,1717..2424)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY"
     primer_bind     complement(1756..1777)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2017..2036)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2364..2385
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2464..2574
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="DEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     polyA_signal    2736..2870
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2773..2792)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2827..2846
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2901..2917)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2901..2917)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2914..2936)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2925..2941
                     /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(2949..2955,2956..2973,2974..2979))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2994..3015
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3132..3149)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3303..3891)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3383..3402)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4062..4853,4854..4922))
                     /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     4685..4704
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4923..5027)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5095..5113
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5151..5173)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5273..5292
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5486..5508
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5500..5517
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5501..5517
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 tcgaggtgag ccccacgttc tgcttcactc tccccatctc ccccccctcc ccacccccaa
       61 ttttgtattt atttattttt taattatttt gtgcagcgat gggggcgggg gggggggggn
      121 cgcgcgccag gcggggcggg gcggggsgrg gggcggggcg gggcgaggcg gagaggtgcg
      181 gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg
      241 cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc
      301 cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta
      361 ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt
      421 taatgacggc tcgtttcttt tctgtggctg cgtgaaagcc ttaaagggct ccgggagggc
      481 cctttgtgcg ggggggagcg gctcgggggg tgcgtgcgtg tgtgtgtgcg tggggagcgc
      541 cgcgtgcggc tccgcgctgc ccggcggctg tgagcgctgc gggcgcggcg cggggctttg
      601 tgcgctccgc agtgtgcgcg aggggagcgc ggccgggggc ggtgccccgc ggtgcggggg
      661 gggctgcgag gggaacaaag gctgcgtgcg gggtgtgtgc gtgggggggt gagcaggggg
      721 tgtgggcgcg gcggtcgggc tgtaaccccc ccctgcaccc ccctccccga gttgctgagc
      781 acggcccggc ttcgggtgcg gggctccgtg cggggcgtgg cgcggggctc gccgtgccgg
      841 gcggggggtg gcggcaggkg ggggtgccgg gcggggcggg gccgcctcgg gccggggagg
      901 gctcggggga ggggcgcggc ggcccccgga gcgccggcgg ctgtcgaggc gcggcgagcc
      961 gcagccattg ccttttatgg taatcgtgcg agagggcgca gggacttcct ttgtcccaaa
     1021 tctgtgcgga gccgaaatct gggaggcgcc gccgcacccc ctctagcggg cgcggggcga
     1081 agcggtgcgg cgccggcagg aaggaaatgg gcggggaggg ccttcgtgcg tcgccgcgcc
     1141 gccgtcccct tctccatctc cagcctcggg gctgtccgca gggggacggc tgccttcggg
     1201 ggggacgggg cagggcgggg ttcggcttct ggcgtgtgac cggcggggtt tatatcttcc
     1261 cttctctgtt cctccgcagc ccccaagctt ggcgcgccta gcgctaccgg actcagatct
     1321 cgagctcaag cttcgaattc aatgtcggct accgctgcca ccgtcccgcc tgccgccccg
     1381 gccggcgagg gtggcccccc tgcacctcct ccaaatctta ccagtaacag gagactgcag
     1441 cagacccagg cccaggtgga tgaggtggtg gacatcatga gggtgaatgt ggacaaggtc
     1501 ctggagcgag accagaagct atcggaactg gatgatcgcg cagatgccct ccaggcaggg
     1561 gcctcccagt ttgaaacaag tgcagccaag ctcaagcgca aatactggtg gaaaaacctc
     1621 aagatgatga tcatcttggg agtgatttgc gccatcatcc tcatcatcat catcgtttac
     1681 ttcagcactg gggatccacc ggtcgccacc atggtgagca agggcgagga gctgttcacc
     1741 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
     1801 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
     1861 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
     1921 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     1981 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     2041 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     2101 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     2161 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
     2221 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     2281 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa
     2341 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     2401 actctcggca tggacgagct gtacggagat ctaggaggat ctggaggcgg ctctggcgga
     2461 ggggacgaga aaaccaccgg ctggagaggc ggccacgtgg tggagggcct ggccggcgag
     2521 ctggagcagc tgagagccag actggagcac caccctcagg gccagagaga gccttcgaat
     2581 tcgtaggata tccagcacag tggcggccgc tcgagtctag ataagtaatg atcataatca
     2641 gccatatcac atctgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga
     2701 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     2761 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     2821 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc cccgggtacc
     2881 gagctcgaat tcgtaatcat gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     2941 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     3001 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     3061 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     3121 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     3181 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     3241 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     3301 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     3361 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     3421 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     3481 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     3541 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     3601 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     3661 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     3721 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     3781 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     3841 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     3901 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     3961 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     4021 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     4081 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     4141 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     4201 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     4261 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     4321 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     4381 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     4441 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     4501 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     4561 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     4621 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     4681 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     4741 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     4801 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     4861 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     4921 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     4981 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     5041 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     5101 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     5161 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     5221 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca
     5281 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
     5341 agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     5401 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
     5461 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc
     5521 aagttgggat ctttgcattg gcccacggct ctcaggatgg ggatgctccc cttcagcacc
     5581 cggttcccct tggaaactga tggtcctggc tctgtggcat ggcagtggca ctgtgaggag
     5641 ccccctacca gcagcacaca gtggggttgg cactgccacg ctccggatgc cgcgctctga
     5701 tccaacccca taatcaaggg aacccgaatt gccccatcat tgcccccacc acccccatcc
     5761 tgccgggccc tcacacccca cgctgccttc tggtgacatt ccccagccca aacccacggc
     5821 ttcatggcta ccgcggggca tttcccattg ccgccccatt atcagctctg cacacctccc
     5881 gctgtaccca tgcctcgtgg ctgcccttct ttgacgtata atcttctaat taatacccgg
     5941 ccttgtcaaa gtggagcaca aacgttaatt aattccccag caggcaggta attaacagtg
     6001 tgactccctt tttgctgcga gtggggctga tacagagaga tgtggcacta tggagcccac
     6061 ggggtcctgg cactgggtgc ccacggaggt ccccatgtgc tgcagtgtca ccgcctccga
     6121 ggagacagta ttgtccctgc ggtgtccctg cagctcagct ctgtccacag ggccacctcc
     6181 agtttggagg ggacacaatg cagccccgat gcaacccatc ctcgcagcat cccagggaca
     6241 aagaccccac tgcaagaccg cacacagggc tgggtcccgc tcccctaata tctacagtgc
     6301 ttcttgcatg gccccttaat caatgcagtt aatcagcatg cgctcatgca ccgctctgga
     6361 gctgcaaagc ccctcgcagc gctgctcacc aacaccgcgc accgccccgg cccagcctgc
     6421 agcacgcgct gcaaacagga aagaaacaaa atattgccca aatgtaggca aaggcattcg
     6481 gctgccttga cctccgccgg gccgggccct gcctgactca gctccttact cagcgctcgc
     6541 ttcctccctc cggctgccac cgccgcagcg cacaccctga caaagagtgg cccttaacgg
     6601 gctctgaggt gcacccagca gtgcactcag cagtccaagg gccggcctgg aggtttgcac
     6661 cgctacgtgc tgacattagc attgaacttg gccctgggta gtgctgcagg ccgggcgggg
     6721 tgggtgtaga gagtgcagcg cgcgttgcac ccggtgcccc ttcccctccc ttgcatccca
     6781 gcaggctgca ccccagcacc aggcccgtgc atgcatgctc ctggtgttat tgcagcctgg
     6841 tgcatgcatg cgtcttgatg gtgcagcgct gtgcatgcat cctccttggt gtgtagcagt
     6901 ctagtgcatg catacccctc ggtgttattg ctgctctgtg cacgcacgct cattgtatca
     6961 cttcatccca gtgcatgcac tcacactgga gcgattgctg ctcggtgcac gcacactcat
     7021 tgtatcactg cagctcagtg gctgcacgca caccggtgtt attgctgctc ggtgcgtgca
     7081 tgcacatcag tgtcgctgca gctcagtgca tgcacgctca ttgcccatcg ctatccctgc
     7141 ctctcctgct ggcgctcccc gggaggtgac ttcaagggga ccgcaggacc acctcggggg
     7201 tggggggagg gctgcacacg cggaccccgc tccccctccc caacaaagca ctgtggaatc
     7261 aaaaaggggg gaggggggat ggaggggcgc gtcacacccc cgccccacac cctcacc
//
