LOCUS       sequence-335635-        7497 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 7497)
  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 7497)
  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..7497
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        45..492
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      511..520
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     517..594
                     /codon_start=1
                     /product="human growth hormone signal sequence"
                     /label=hGH signal sequence
                     /translation="MATGSRTSLLLAFGLLCLPWLQEGSA"
     CDS             601..1074
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="ISLIAALAVDRVIGMENAMPWNLPADLAWFKRNTLNKPVIMGRHT
                     WESIGRPLPGRKNIILSSQPGTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFLP
                     KAQKLYLTHIDAEVEGDTHFPDYEPDDWESVFSEFHDADAQNSHSYCFEILERR"
     CDS             join(1081..1083,1084..1794)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1081..1083,1084..1794)
                     /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(1123..1144)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1384..1403)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1731..1752
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1795..1812
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1825..1851
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2247..2256
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    4285..4406
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4322..4341)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4376..4395
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4413..4868)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4550..4571)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4762..4781
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4895..4999
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5001..5358
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5001..5192
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5024..5044)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5209..5344
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5393..6187
                     /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             5393..6187
                     /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(5447..5466)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6057..6076
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6375..6394)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6419..6466
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6795..7383
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7284..7303
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa ttaagtgtct agactgcaga gggccctgcg tatgagtgca agtgggtttt
       61 aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga cccactggac
      121 aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg gaggggaaac
      181 aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt gccttcgccc
      241 ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac gtcactcgcc
      301 ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc cgccggccca
      361 gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca tctgcgctgc
      421 ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt cgtgtcgtgc
      481 ctgagagcgc agtcgagaaa ccggtctcga gccaccatgg ctacaggctc ccggacgtcc
      541 ctgctcctgg cttttggcct gctctgcctg ccctggcttc aagagggcag tgccgagctc
      601 atcagtctga ttgcggcgtt agcggtagat cgcgttatcg gcatggaaaa cgccatgccg
      661 tggaacctgc ctgccgatct cgcctggttt aaacgcaaca ccttaaataa acccgtgatt
      721 atgggccgcc atacctggga atcaatcggt cgtccgttgc caggacgcaa aaatattatc
      781 ctcagcagtc aaccgggtac ggacgatcgc gtaacgtggg tgaagtcggt ggatgaagcc
      841 atcgcggcgt gtggtgacgt accagaaatc atggtgattg gcggcggtcg cgtttatgaa
      901 cagttcttgc caaaagcgca aaaactgtat ctgacgcata tcgacgcaga agtggaaggc
      961 gacacccatt tcccggatta cgagccggat gactgggaat cggtattcag cgaattccac
     1021 gatgctgatg cgcagaactc tcacagctat tgctttgaga ttctggagcg gcgggtcgac
     1081 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     1141 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     1201 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1261 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     1321 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     1381 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     1441 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     1501 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     1561 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     1621 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     1681 ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     1741 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagcttgtt
     1801 cctcgaggaa gcaagcttca aaagtatcca tacgacgttc cggactacgc attggatgat
     1861 gtagacccat tggttactac taactttggc aagattaggg gaattaagaa agaactcaat
     1921 aatgaaattt tgggtcctgt cattcagttt cttggggttc catatgccgc tccaccaaca
     1981 ggagaacatc gtttccagcc tccagaacca ccatctccct ggtctgacat ccggaacgcc
     2041 actcagtttg ctcctgtatg tccccagaat atcattgatg gcagattgcc tgaagttatg
     2101 cttcctgtgt ggttcactaa taacttggat gtggtttcat catacgtcca agaccagagt
     2161 gaagactgtc tatacttaaa catctatgtc ccaactgaag atgatattcg ggacagtggg
     2221 ggtcccaaac cagtgatggt gtacatccat ggcggctctt acatggaagg tactggaaat
     2281 ctgtatgatg ggagtgtctt ggcaagctat ggcaatgtga tcgtcatcac agtcaactat
     2341 cggcttgggg tacttggctt cttgagcaca ggggatcagg ctgccaaagg aaactacggg
     2401 ctccttgacc tcatccaggc cctaagatgg accagcgaga acattgggtt ctttggtggt
     2461 gaccccttgc gaatcactgt gtttggatca ggcgctgggg gttcatgtgt caacctgctg
     2521 actttatccc attattctga aggacttttt caacgagcaa tagctcagag tggaacagcc
     2581 ctttccagct gggctgttag tttccagcct gcaaaatacg ctagaattct ggccacaaaa
     2641 gttggctgca atgtttcaga tacagtagag ttagtagaat gcctgcagaa gaagccttac
     2701 aaagaacttg ttgatcaaga tgttcaacca gcccgatacc acatagcctt tggacctgtg
     2761 atcgatggtg atgtaatacc agatgaccct cagatactga tggaacaagg agagttcctc
     2821 aactatgata taatgttggg agttaaccaa ggggaagggt tgaagtttgt cgaaaatata
     2881 gtagatagtg atgatggtgt atcagccagt gatttcgact ttgctgtttc taattttgtt
     2941 gataatttat atggatatcc tgaaggcaaa gatgttttga gagaaaccat taaattcatg
     3001 tatactgact gggctgatcg ccataatcct gaaactagaa ggaagacatt gttggctttg
     3061 tttacggacc atcaatgggt agcacctgct gtggccacag cagaccttca ctcgaacttt
     3121 ggctcaccta catacttcta tgccttttat catcattgcc aaacagatca agttccagct
     3181 tgggctgatg cagctcatgg ggatgaggtt ccctatgtgt tgggaatccc catgattggc
     3241 cctacagagt tatttccttg caatttctcc aagaatgatg tgatgttgag tgcagtagta
     3301 atgacatact ggacgaattt tgctaaaact ggtgacccaa atcaaccagt tcctcaagac
     3361 acaaaattca tccataccaa acccaaccgc tttgaagaag tagcatggac cagatattcc
     3421 cagaaagacc agctttatct ccatattgga ttaaaaccga gagttaaaga gcattacaga
     3481 gccaataagg taaatctctg gctggagctg gtacctcatc tgcataatct caatgacatt
     3541 tctcagtata cctcgacaac aactaaagtg ccatccacgg acatcactct cagacctaca
     3601 aggaagaatt ccacaccagt cacatcagcc tttcccactg ccaaacagga tgatcccaag
     3661 caacaaccaa gccccttctc ggtggatcag agggactact ccacagagct aagtgtcact
     3721 atcgcagtgg gggcctctct gctgtttctc aacatcttgg cttttgcagc cctgtactac
     3781 aagaaggata agaggagaca tgatgtccac cggaggtgca gccctcagcg cacgaccacc
     3841 aacgacctaa cccatgctcc agaagaggaa attatgtctc tccaaatgaa gcacactgac
     3901 ttggatcacg agtgtgagtc catccatcca catgaggtgg ttcttcggac cgcctgtccc
     3961 ccagattata ctctagctat gaggaggtca cctgatgata ttccactaat gacacctaac
     4021 accatcacaa tgattcccaa cactatacca gggattcagc ccttacatac attcaacaca
     4081 tttactggag gacagaataa tacactgccc catccccacc cacaccccca ttcacattca
     4141 acaaccaggg tatagggatc caccggatct agataactga tcataatcag ccataccaca
     4201 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     4261 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     4321 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     4381 ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa
     4441 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     4501 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     4561 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     4621 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     4681 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     4741 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     4801 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     4861 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     4921 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     4981 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     5041 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     5101 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     5161 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     5221 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     5281 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     5341 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
     5401 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     5461 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     5521 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     5581 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     5641 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     5701 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     5761 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     5821 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     5881 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     5941 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     6001 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     6061 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     6121 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     6181 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     6241 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     6301 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
     6361 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     6421 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     6481 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     6541 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     6601 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     6661 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     6721 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     6781 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     6841 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     6901 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     6961 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     7021 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     7081 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     7141 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     7201 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     7261 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     7321 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     7381 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     7441 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
