LOCUS       sequence-339111-        7494 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For light regulated release of neuroligin 1 from the endoplasmic 
            reticulum. Driven by synapsin promoter. Thrombin site is included 
            between GFP and NL1..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7494)
  AUTHORS   Bourke AM, Schwartz SL, Bowen AB, Kleinjan MS, Winborn CS, Kareemo 
            DJ, Gutnick A, Schwarz TL, Kennedy MJ
  TITLE     zapERtrap: A light-regulated ER release system reveals unexpected 
            neuronal trafficking pathways.
  JOURNAL   J Cell Biol. 2021 Sep 6;220(9). pii: 212461. doi: 
            10.1083/jcb.202103186. Epub 2021 Jul 9.
  PUBMED    34241635
REFERENCE   2  (bases 1 to 7494)
  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: "J Cell 
            Biol."; date: "2021-09-6"; volume: "220(9). pii: 212461. doi"; 
            pages: " 10.1083/jcb.202103186. Epub 2021 Jul 9"
COMMENT     Sequence Label: DHFR-GFP-th-NL1
FEATURES             Location/Qualifiers
     source          1..7494
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..468
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      487..496
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     493..570
                     /codon_start=1
                     /product="human growth hormone signal sequence"
                     /label=hGH signal sequence
                     /translation="MATGSRTSLLLAFGLLCLPWLQEGSA"
     CDS             577..1050
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="ISLIAALAVDRVIGMENAMPWNLPADLAWFKRNTLNKPVIMGRHT
                     WESIGRPLPGRKNIILSSQPGTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFLP
                     KAQKLYLTHIDAEVEGDTHFPDYEPDDWESVFSEFHDADAQNSHSYCFEILERR"
     CDS             join(1057..1059,1060..1770)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1057..1059,1060..1770)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1099..1120)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1360..1379)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1707..1728
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1771..1788
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1801..1827
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2223..2232
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    4261..4382
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4298..4317)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4352..4371
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4389..4844)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4526..4547)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4738..4757
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4871..4975
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4977..5334
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4977..5168
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5000..5020)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5185..5320
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5369..6163
                     /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             5369..6163
                     /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(5423..5442)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6033..6052
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6351..6370)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6395..6442
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6771..7359
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7260..7279
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgcagagggc cctgcgtatg agtgcaagtg ggttttagga ccaggatgag gcggggtggg
       61 ggtgcctacc tgacgaccga ccccgaccca ctggacaagc acccaacccc cattccccaa
      121 attgcgcatc ccctatcaga gagggggagg ggaaacagga tgcggcgagg cgcgtgcgca
      181 ctgccagctt cagcaccgcg gacagtgcct tcgcccccgc ctggcggcgc gcgccaccgc
      241 cgcctcagca ctgaaggcgc gctgacgtca ctcgccggtc ccccgcaaac tccccttccc
      301 ggccaccttg gtcgcgtccg cgccgccgcc ggcccagccg gaccgcacca cgcgaggcgc
      361 gagatagggg ggcacgggcg cgaccatctg cgctgcggcg ccggcgactc agcgctgcct
      421 cagtctgcgg tgggcagcgg aggagtcgtg tcgtgcctga gagcgcagtc gagaaaccgg
      481 tctcgagcca ccatggctac aggctcccgg acgtccctgc tcctggcttt tggcctgctc
      541 tgcctgccct ggcttcaaga gggcagtgcc gagctcatca gtctgattgc ggcgttagcg
      601 gtagatcgcg ttatcggcat ggaaaacgcc atgccgtgga acctgcctgc cgatctcgcc
      661 tggtttaaac gcaacacctt aaataaaccc gtgattatgg gccgccatac ctgggaatca
      721 atcggtcgtc cgttgccagg acgcaaaaat attatcctca gcagtcaacc gggtacggac
      781 gatcgcgtaa cgtgggtgaa gtcggtggat gaagccatcg cggcgtgtgg tgacgtacca
      841 gaaatcatgg tgattggcgg cggtcgcgtt tatgaacagt tcttgccaaa agcgcaaaaa
      901 ctgtatctga cgcatatcga cgcagaagtg gaaggcgaca cccatttccc ggattacgag
      961 ccggatgact gggaatcggt attcagcgaa ttccacgatg ctgatgcaca gaactctcac
     1021 agctattgct ttgagattct ggagcggcgg gtcgacgtga gcaagggcga ggagctgttc
     1081 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     1141 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     1201 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     1261 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     1321 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     1381 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     1441 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
     1501 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
     1561 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     1621 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
     1681 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     1741 atcactctcg gcatggacga gctgtacaag cttgttcctc gaggaagcaa gcttcaaaag
     1801 tatccatacg acgttccgga ctacgcattg gatgatgtag acccattggt tactactaac
     1861 tttggcaaga ttaggggaat taagaaagaa ctcaataatg aaattttggg tcctgtcatt
     1921 cagtttcttg gggttccata tgccgctcca ccaacaggag aacatcgttt ccagcctcca
     1981 gaaccaccat ctccctggtc tgacatccgg aacgccactc agtttgctcc tgtatgtccc
     2041 cagaatatca ttgatggcag attgcctgaa gttatgcttc ctgtgtggtt cactaataac
     2101 ttggatgtgg tttcatcata cgtccaagac cagagtgaag actgtctata cttaaacatc
     2161 tatgtcccaa ctgaagatga tattcgggac agtgggggtc ccaaaccagt gatggtgtac
     2221 atccatggcg gctcttacat ggaaggtact ggaaatctgt atgatgggag tgtcttggca
     2281 agctatggca atgtgatcgt catcacagtc aactatcggc ttggggtact tggcttcttg
     2341 agcacagggg atcaggctgc caaaggaaac tacgggctcc ttgacctcat ccaggcccta
     2401 agatggacca gcgagaacat tgggttcttt ggtggtgacc ccttgcgaat cactgtgttt
     2461 ggatcaggcg ctgggggttc atgtgtcaac ctgctgactt tatcccatta ttctgaagga
     2521 ctttttcaac gagcaatagc tcagagtgga acagcccttt ccagctgggc tgttagtttc
     2581 cagcctgcaa aatacgctag aattctggcc acaaaagttg gctgcaatgt ttcagataca
     2641 gtagagttag tagaatgcct gcagaagaag ccttacaaag aacttgttga tcaagatgtt
     2701 caaccagccc gataccacat agcctttgga cctgtgatcg atggtgatgt aataccagat
     2761 gaccctcaga tactgatgga acaaggagag ttcctcaact atgatataat gttgggagtt
     2821 aaccaagggg aagggttgaa gtttgtcgaa aatatagtag atagtgatga tggtgtatca
     2881 gccagtgatt tcgactttgc tgtttctaat tttgttgata atttatatgg atatcctgaa
     2941 ggcaaagatg ttttgagaga aaccattaaa ttcatgtata ctgactgggc tgatcgccat
     3001 aatcctgaaa ctagaaggaa gacattgttg gctttgttta cggaccatca atgggtagca
     3061 cctgctgtgg ccacagcaga ccttcactcg aactttggct cacctacata cttctatgcc
     3121 ttttatcatc attgccaaac agatcaagtt ccagcttggg ctgatgcagc tcatggggat
     3181 gaggttccct atgtgttggg aatccccatg attggcccta cagagttatt tccttgcaat
     3241 ttctccaaga atgatgtgat gttgagtgca gtagtaatga catactggac gaattttgct
     3301 aaaactggtg acccaaatca accagttcct caagacacaa aattcatcca taccaaaccc
     3361 aaccgctttg aagaagtagc atggaccaga tattcccaga aagaccagct ttatctccat
     3421 attggattaa aaccgagagt taaagagcat tacagagcca ataaggtaaa tctctggctg
     3481 gagctggtac ctcatctgca taatctcaat gacatttctc agtatacctc gacaacaact
     3541 aaagtgccat ccacggacat cactctcaga cctacaagga agaattccac accagtcaca
     3601 tcagcctttc ccactgccaa acaggatgat cccaagcaac aaccaagccc cttctcggtg
     3661 gatcagaggg actactccac agagctaagt gtcactatcg cagtgggggc ctctctgctg
     3721 tttctcaaca tcttggcttt tgcagccctg tactacaaga aggataagag gagacatgat
     3781 gtccaccgga ggtgcagccc tcagcgcacg accaccaacg acctaaccca tgctccagaa
     3841 gaggaaatta tgtctctcca aatgaagcac actgacttgg atcacgagtg tgagtccatc
     3901 catccacatg aggtggttct tcggaccgcc tgtcccccag attatactct agctatgagg
     3961 aggtcacctg atgatattcc actaatgaca cctaacacca tcacaatgat tcccaacact
     4021 ataccaggga ttcagccctt acatacattc aacacattta ctggaggaca gaataataca
     4081 ctgccccatc cccacccaca cccccattca cattcaacaa ccagggtata gggatccacc
     4141 ggatctagat aactgatcat aatcagccat accacatttg tagaggtttt acttgcttta
     4201 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt
     4261 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     4321 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     4381 taacgcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     4441 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     4501 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     4561 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     4621 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     4681 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     4741 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     4801 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg
     4861 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     4921 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga
     4981 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     5041 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     5101 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     5161 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     5221 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     5281 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga
     5341 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     5401 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     5461 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     5521 cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc
     5581 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     5641 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     5701 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     5761 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     5821 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     5881 cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     5941 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     6001 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     6061 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     6121 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     6181 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     6241 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     6301 cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg
     6361 aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg
     6421 cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc
     6481 gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc
     6541 caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct
     6601 gccatagcct caggttactc atatatactt tagattgatt taaaacttca tttttaattt
     6661 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     6721 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     6781 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     6841 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     6901 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     6961 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     7021 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     7081 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     7141 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     7201 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     7261 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     7321 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     7381 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     7441 gattctgtgg ataaccgtat taccgccatg cattagttat ttaagtgtct agac
//
