LOCUS       Exported                7010 bp ds-DNA     circular SYN 30-AUG-2023
DEFINITION  Global labeling of excitatory synapses (green fluorescence).
ACCESSION   .
VERSION     .
KEYWORDS    AAV-EF1A-PSD95.FingR-eGFP-CCR5TC
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7010)
  AUTHORS   Bensussen S, Shankar S, Ching KH, Zemel D, Ta TL, Mount RA, Shroff 
            SN, Gritton HJ, Fabris P, Vanbenschoten H, Beck C, Man HY, Han X
  TITLE     A Viral Toolbox of Genetically Encoded Fluorescent Synaptic Tags.
  JOURNAL   iScience. 2020 Jun 30;23(7):101330. doi: 10.1016/j.isci.2020.101330.
  PUBMED    32674057
REFERENCE   2  (bases 1 to 7010)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 30, 2023 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7010
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     94..111
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    228..249
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        264..294
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    302..318
                     /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     307..329
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     326..342
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     326..342
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   380..509
                     /label=AAV2 ITR
     protein_bind    621..632
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     promoter        644..1822
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          875..1813
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1770..1790
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1839..1858
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1839..1857
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2181..2204
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2229..2939
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELY"
     CDS             2229..2939
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELY"
     primer_bind     complement(2271..2292)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2532..2551)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2879..2900
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2985..3326
                     /codon_start=1
                     /product="zinc-finger domain from the CCR5-224 (+) 
                     zinc-finger nuclease that recognizes the left half-site 
                     target sequence in the human CCR5 gene (Perez et al., 2008;
                     Gross et al., 2013)"
                     /label=CCR5TC
                     /translation="FQCRICMRNFSDRSNLSRHIRTHTGEKPFACDICGRKFAISSNLN
                     SHTKIHTGSQKPFQCRICMRNFSRSDNLARHIRTHTGEKPFACDICGRKFATSGNLTRH
                     TKIHLRGSQL"
     CDS             3375..3500
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the rat zinc finger protein Kid-1 
                     (Witzgall et al., 1994)"
                     /label=KRAB-A
                     /translation="VSVTFEDVAVLFTRDEWKKLDLSQRSLYREVMLENYSNLASM"
     misc_feature    3519..4107
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3572..3592)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3990..4001)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4110..4126)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4111..4127)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    4156..4277
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4167..4186)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4221..4240
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4439..4568
                     /label=AAV2 ITR
     primer_bind     complement(4587..4604)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4587..4603)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4596..4618)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4745..5200
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4832..4851)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5042..5063
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5226..5330
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5331..6191
                     /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(5549..5568)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6362..6950
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6851..6870
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
       61 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      121 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
      181 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
      241 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
      301 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cgccagattt
      361 aattaaggcc ttaattaggc tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc
      421 cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg
      481 agtggccaac tccatcacta ggggttcctt gtagttaatg attaacccgc catgctactt
      541 atctacgtag ccatgctcta ggaagatctg gacagcagag atccagtttg gatcgataag
      601 cttgatatcg aattacgcgt gtcatcctca tcgaattccg tgaggctccg gtgcccgtca
      661 gtgggcagag cgcacatcgc ccacagtccc cgagaagttg gggggagggg tcggcaattg
      721 aaccggtgcc tagagaaggt ggcgcggggt aaactgggaa agtgatgtcg tgtactggct
      781 ccgccttttt cccgagggtg ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt
      841 tctttttcgc aacgggtttg ccgccagaac acaggtaagt gccgtgtgtg gttcccgcgg
      901 gcctggcctc tttacgggtt atggcccttg cgtgccttga attacttcca cctggctgca
      961 gtacgtgatt cttgatcccg agcttcgggt tggaagtggg tgggagagtt cgaggccttg
     1021 cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc tggcctgggc gctggggccg
     1081 ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct gctttcgata agtctctagc
     1141 catttaaaat ttttgatgac ctgctgcgac gctttttttc tggcaagata gtcttgtaaa
     1201 tgcgggccaa gatctgcaca ctggtatttc ggtttttggg gccgcgggcg gcgacggggc
     1261 ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg cgagcgcggc caccgagaat
     1321 cggacggggg tagtctcaag ctggccggcc tgctctggtg cctggcctcg cgccgccgtg
     1381 tatcgccccg ccctgggcgg caaggctggc ccggtcggca ccagttgcgt gagcggaaag
     1441 atggccgctt cccggccctg ctgcagggag ctcaaaatgg aggacgcggc gctcgggaga
     1501 gcgggcgggt gagtcaccca cacaaaggaa aagggccttt ccgtcctcag ccgtcgcttc
     1561 atgtgactcc acggagtacc gggcgccgtc caggcacctc gattagttct cgagcttttg
     1621 gagtacgtcg tctttaggtt ggggggaggg gttttatgcg atggagtttc cccacactga
     1681 gtgggtggag actgaagtta ggccagcttg gcacttgatg taattctcct tggaatttgc
     1741 cctttttgag tttggatctt ggttcattct caagcctcag acagtggttc aaagtttttt
     1801 tcttccattt caggtgtcgt gaggaattag cttggtacta atacgactca ctatagggag
     1861 acccaagctg gctaggtaag cttggtaccg agctcggatc cgccgccacc atgctcgaag
     1921 tcaaggaagc atcaccaacc agcatccaga tcagctgggt gctccacttg cgccacgttc
     1981 gctactaccg catcacctac ggtgaaactg gtggcaatag ccctgtccag gaattcaccg
     2041 tgcctggcag caagtccact gctaccatca gcggcctgaa acctggtgtc gactatacca
     2101 tcacggtgta cgccgtcacg atcttcagcg cctaccgctc cgcctggccg ccgatctcca
     2161 tcaactaccg caccggaacc gattacaagg atgacgacga taagggtagc ggctccagta
     2221 gatctggggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2281 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2341 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2401 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     2461 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     2521 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     2581 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     2641 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     2701 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     2761 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     2821 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     2881 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtact
     2941 ctaggggagc tggcgctgga gcgggtgcag gggctggctc tagattccag tgccggatct
     3001 gcatgcggaa cttcagcgac cggtccaacc tgagcaggca catcagaacc cacaccggag
     3061 aaaagccctt cgcctgcgac atttgcggcc ggaagttcgc catcagcagc aacctgaaca
     3121 gccacaccaa gatccacact ggcagccaga aacctttcca gtgcagaatt tgtatgagaa
     3181 actttagcag aagcgacaac ctggccagac acatccggac acatactggt gaaaaacctt
     3241 ttgcctgtga tatctgtggc agaaagtttg ccacctccgg caatctgacc cggcacacaa
     3301 agattcacct gcggggcagc cagctatcga ttgtcgacgc tcctgaacaa cgtgaaggtg
     3361 cttctcaagt ttctgtttct gttacttttg aagatgttgc tgttcttttt actcgtgatg
     3421 aatggaaaaa acttgatctt tctcaacgtt ctctttatcg tgaagttatg cttgaaaatt
     3481 attctaatct tgcttctatg gcttaaaagc ttatcgataa tcaacctctg gattacaaaa
     3541 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     3601 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     3661 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     3721 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     3781 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     3841 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     3901 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc
     3961 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     4021 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     4081 ggatctccct ttgggccgcc tccccgcatc gataccgtcg acctcgaccc gggcggccgc
     4141 ttcgagcaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     4201 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     4261 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     4321 agatgtggga ggttttttaa agcaagtaaa acctctacaa atgtggtaaa atcgataagg
     4381 atcttcctag agcatggcta cgtagataag tagcatggcg ggttaatcat taactacaag
     4441 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc
     4501 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga
     4561 gcgcgcagcc ttaattaacc taattcactg gccgtcgttt tacaacgtcg tgactgggaa
     4621 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
     4681 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     4741 tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     4801 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     4861 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     4921 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     4981 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     5041 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     5101 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     5161 tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa
     5221 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     5281 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     5341 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     5401 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     5461 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     5521 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     5581 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     5641 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     5701 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     5761 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5821 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     5881 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5941 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     6001 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     6061 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     6121 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     6181 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     6241 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     6301 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     6361 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     6421 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     6481 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     6541 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     6601 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     6661 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     6721 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     6781 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     6841 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     6901 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     6961 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
//
