LOCUS       Exported                5943 bp DNA     circular SYN 14-OCT-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5943)
  AUTHORS   Fishell Lab
  TITLE     Direct Submission
  JOURNAL   Exported Nov 3, 2021 from SnapGene 5.3.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5943
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     misc_feature    1..130
                     /label=ITR
     misc_feature    150..608
                     /label=hSyn1 Promoter
     misc_feature    609..628
                     /label=P/E cut
     misc_feature    609..622
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     misc_feature    609..622
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     misc_feature    623..628
                     /label=Gibs_Flex_Fb
     misc_feature    623..628
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    629..662
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     protein_bind    629..662
                     /label=LoxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    651..662
                     /label=Gibson_Flex_R1
     protein_bind    663..696
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    747..780
                     /label=F5
     misc_feature    781..782
                     /label=Changed gg to aa to remove AscI site
     misc_feature    792..825
                     /label=MCS
                     /note=" multiple cloning site
                     "
     primer_bind     810..852
                     /label=GA-GCaMP6f.Rev
     misc_feature    830..2189
                     /label=Insert_2
     CDS             complement(834..2183)
                     /codon_start=1
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     1793..1812
                     /label=EGFP.R
                     /note="20 mer	60"
     primer_bind     complement(1851..1868)
                     /label=EGFP.F
                     /note="18 mer	61"
     primer_bind     complement(2170..2209)
                     /label=GA-GCaMP6f.For
     misc_feature    2184..2189
                     /label=Kozak
     misc_feature    2195..2213
                     /label=MCS
                     /note=" multiple cloning site
                     "
     protein_bind    complement(2224..2257)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    complement(2308..2341)
                     /label=F5
     misc_feature    2342..2375
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     protein_bind    2342..2375
                     /label=LoxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    2364..2375
                     /label=Gibson_Flex_R1
     misc_feature    2376..2383
                     /label=AscI
     misc_feature    2384..2972
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     polyA_signal    2980..3204
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3206..3346
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      3421..3876
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4158..4262
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4263..5123
                     /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"
     rep_origin      5294..5882
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gctagactgc agagggccct gcgtatgagt gcaagtgggt tttaggacca
      181 ggatgaggcg gggtgggggt gcctacctga cgaccgaccc cgacccactg gacaagcacc
      241 caacccccat tccccaaatt gcgcatcccc tatcagagag ggggagggga aacaggatgc
      301 ggcgaggcgc gtgcgcactg ccagcttcag caccgcggac agtgccttcg cccccgcctg
      361 gcggcgcgcg ccaccgccgc ctcagcactg aaggcgcgct gacgtcactc gccggtcccc
      421 cgcaaactcc ccttcccggc caccttggtc gcgtccgcgc cgccgccggc ccagccggac
      481 cgcaccacgc gaggcgcgag ataggggggc acgggcgcga ccatctgcgc tgcggcgccg
      541 gcgactcagc gctgcctcag tctgcggtgg gcagcggagg agtcgtgtcg tgcctgagag
      601 cgcagtcaga tatcgtttaa actccggaat aacttcgtat agcatacatt atacgaagtt
      661 atgaagttcc tattctctag aaagtatagg aacttcgcag aatggtagct ggattgtagc
      721 tgctattagc aatatgaaac ctcttagaag ttcctattct tcaaaaggta taggaacttc
      781 aacgcgccct agaattcgag ctcggtaccc ggggatcctc tagagtcgac tcacttcgct
      841 gtcatcattt gtacaaactc ttcgtagttt acctgaccat ccccatcgat gtctgcttcc
      901 ctgatcattt catcaacctc ttcatctgtt aacttctctc caaggtttgt catcacgtgg
      961 cgaagctctg ctgcactgat gtagccattg ccatccttat caaacacacc gaacgcttct
     1021 ctaatttctt cttccgtgtc cctgtatttc atttttcttg ccatcattgt caggaactca
     1081 gggaagtcga ttgtgccgtc accgtcggca tctacttcat tgatcatgtc ctgcagctct
     1141 gcttctgtgg ggttctgccc cagagaccgc atcaccgtcc ccagctcctt ggttgttatt
     1201 gtcccatccc cgtccttgtc aaatagggag aattcctctt taaattctgc gatctgctct
     1261 tcagtcagtt ggtccggcag gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc
     1321 ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc
     1381 tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc ctggatgtag
     1441 ccttcgggca tggcggactt gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg
     1501 aagcactgca cgccgtaggt cagggtggtc acgagggtgg gccagggcac gggcagcttg
     1561 ccggtggtgc agatgaactt cagggtcagc ttgccgtagg tggcatcgcc ctcaccctcg
     1621 ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag gatgggcacc
     1681 accccggtga acagctcctc gcccttgctc accatgctcc ctccggtacc gcccttgtac
     1741 agctcgtcca tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga
     1801 tcgcgcttct cgttggggtc tttcgaaagt ttggactgca cgctcaggta gtggttgtcg
     1861 ggcagcagca cggggccgtc gccgatgggg gtgttctgct ggtagtggta ggcgagctgc
     1921 acgccgccgt cctcgatgtt gtggcggatc ttgaagttcg ccttgatgcc gttcttctgc
     1981 ttgtcggcct tgatatagac gttctcgagt gagctcagcc gacctatagc tctgactgcg
     2041 tgacctgtct tattccactt acgacgtgat gagtcgacca tggtggcgag atccttatcg
     2101 tcatcgtcgt acagatcccg acccatttgc tgtccaccag tcatgctagc cataccatga
     2161 tgatgatgat gatgagaacc catggtggcg tcgacctgca ggcatgcaag cttggtggct
     2221 agcgaagttc ctatactttc tagagaatag gaacttcttg ccttaaccca gaaattatca
     2281 ctgttattct ttagaatggt gcaaagagaa gttcctatac cttttgaaga ataggaactt
     2341 cataacttcg tatagcatac attatacgaa gttatggcgc gccaatcaac ctctggatta
     2401 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     2461 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     2521 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     2581 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     2641 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     2701 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     2761 cgtggtgttg tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg
     2821 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     2881 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     2941 gagtcggatc tccctttggg ccgcctcccc gcctggatcc tgtgccttct agttgccagc
     3001 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     3061 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     3121 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     3181 ctggggatgc ggtgggctct atggcaggaa cccctagtga tggagttggc cactccctct
     3241 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     3301 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg
     3361 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     3421 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     3481 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     3541 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     3601 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     3661 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     3721 gactcttgtt ccaaactgga acaacactca accctatctc gggctattct tttgatttat
     3781 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     3841 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     3901 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     3961 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     4021 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     4081 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     4141 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4201 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4261 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     4321 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     4381 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     4441 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     4501 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     4561 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     4621 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     4681 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     4741 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     4801 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     4861 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     4921 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     4981 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     5041 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     5101 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     5161 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     5221 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     5281 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     5341 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     5401 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     5461 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     5521 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     5581 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     5641 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     5701 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     5761 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     5821 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     5881 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     5941 tgt
//
