LOCUS Exported 6238 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Global labeling of excitatory synapses (green fluorescence). ACCESSION . VERSION . KEYWORDS AAV-Syn-PSD95.FingR-eGFP-CCR5TC SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6238) 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 6238) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6238 /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 575..586 /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 614..1061 /label=hSyn promoter /note="human synapsin I promoter; confers neuron-specific expression (Kugler et al., 2003)" CDS 1368..1391 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1416..2126 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELY" CDS 1416..2126 /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(1458..1479) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1719..1738) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2066..2087 /label=EGFP-C /note="EGFP, forward primer" CDS 2172..2513 /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 2562..2687 /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 2747..3335 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2800..2820) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3218..3229) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(3338..3354) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3339..3355) /label=pBluescriptKS /note="For pBluescript vector" polyA_signal 3384..3505 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3395..3414) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3449..3468 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 3667..3796 /label=AAV2 ITR primer_bind complement(3815..3832) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(3815..3831) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3824..3846) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 3973..4428 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4060..4079) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4270..4291 /label=F1ori-F /note="F1 origin, forward primer" promoter 4454..4558 /gene="bla" /label=AmpR promoter CDS 4559..5419 /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(4777..4796) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5590..6178 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6079..6098 /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 ggaagatcac gcgtgtcatc ctcatcagat ctctgcagag 601 ggccctgcgt atgagtgcaa gtgggtttta ggaccaggat gaggcggggt gggggtgcct 661 acctgacgac cgaccccgac ccactggaca agcacccaac ccccattccc caaattgcgc 721 atcccctatc agagaggggg aggggaaaca ggatgcggcg aggcgcgtgc gcactgccag 781 cttcagcacc gcggacagtg ccttcgcccc cgcctggcgg cgcgcgccac cgccgcctca 841 gcactgaagg cgcgctgacg tcactcgccg gtcccccgca aactcccctt cccggccacc 901 ttggtcgcgt ccgcgccgcc gccggcccag ccggaccgca ccacgcgagg cgcgagatag 961 gggggcacgg gcgcgaccat ctgcgctgcg gcgccggcga ctcagcgctg cctcagtctg 1021 cggtgggcag cggaggagtc gtgtcgtgcc tgagagcgca gtcgaattca agctgctagc 1081 aaggatccgc cgccaccatg ctcgaagtca aggaagcatc accaaccagc atccagatca 1141 gctgggtgct ccacttgcgc cacgttcgct actaccgcat cacctacggt gaaactggtg 1201 gcaatagccc tgtccaggaa ttcaccgtgc ctggcagcaa gtccactgct accatcagcg 1261 gcctgaaacc tggtgtcgac tataccatca cggtgtacgc cgtcacgatc ttcagcgcct 1321 accgctccgc ctggccgccg atctccatca actaccgcac cggaaccgat tacaaggatg 1381 acgacgataa gggtagcggc tccagtagat ctggggtgag caagggcgag gagctgttca 1441 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg 1501 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca 1561 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc 1621 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc 1681 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc 1741 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg 1801 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca 1861 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc 1921 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg 1981 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca 2041 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga 2101 tcactctcgg catggacgag ctgtactcta ggggagctgg cgctggagcg ggtgcagggg 2161 ctggctctag attccagtgc cggatctgca tgcggaactt cagcgaccgg tccaacctga 2221 gcaggcacat cagaacccac accggagaaa agcccttcgc ctgcgacatt tgcggccgga 2281 agttcgccat cagcagcaac ctgaacagcc acaccaagat ccacactggc agccagaaac 2341 ctttccagtg cagaatttgt atgagaaact ttagcagaag cgacaacctg gccagacaca 2401 tccggacaca tactggtgaa aaaccttttg cctgtgatat ctgtggcaga aagtttgcca 2461 cctccggcaa tctgacccgg cacacaaaga ttcacctgcg gggcagccag ctatcgattg 2521 tcgacgctcc tgaacaacgt gaaggtgctt ctcaagtttc tgtttctgtt acttttgaag 2581 atgttgctgt tctttttact cgtgatgaat ggaaaaaact tgatctttct caacgttctc 2641 tttatcgtga agttatgctt gaaaattatt ctaatcttgc ttctatggct taaaagcttg 2701 cggccgcgac tctagagtcg acctgcaggc atgcaagctt atcgataatc aacctctgga 2761 ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg 2821 tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt 2881 ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag 2941 gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc 3001 caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga 3061 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa 3121 ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac 3181 ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct 3241 tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca 3301 gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgtcgac ctcgacccgg 3361 gcggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag 3421 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac 3481 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt 3541 tcagggggag atgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat 3601 cgataaggat cttcctagag catggctacg tagataagta gcatggcggg ttaatcatta 3661 actacaagga acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca 3721 ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga 3781 gcgagcgagc gcgcagcctt aattaaccta attcactggc cgtcgtttta caacgtcgtg 3841 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca 3901 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga 3961 atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc 4021 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt 4081 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag 4141 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt 4201 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt 4261 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt 4321 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt 4381 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt 4441 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta 4501 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat 4561 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt 4621 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg 4681 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga 4741 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg 4801 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt 4861 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg 4921 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg 4981 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga 5041 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc 5101 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc 5161 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc 5221 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg 5281 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac 5341 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc 5401 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt 5461 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac 5521 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa 5581 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc 5641 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt 5701 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg 5761 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc 5821 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt 5881 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga 5941 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct 6001 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg 6061 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca 6121 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa 6181 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatg //