LOCUS       Exported                5747 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 5747)
  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..5747
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     misc_feature    1..130
                     /label=ITR
     misc_feature    144..149
                     /label=ApaI
     misc_feature    150..1041
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     primer_bind     151..188
                     /label=Human_Dlx_F_Gibson
     misc_feature    151..166
                     /label=Gibson_hDlx_P_F
     misc_feature    166
                     /label=""A"" after PacI digest
     misc_feature    167..1005
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     misc_feature    167..707
                     /label=hDlx
     misc_feature    167..370
                     /label=h56ii_enhancer
     misc_feature    167..188
                     /label=Gibson_hDlx_I_F
     misc_feature    complement(336..621)
                     /label=hI56i from Zerucha et al., 2000 in reverse
                     /note="100% identical to mouse homologous seq85%identical 
                     to zebrafish homologous seq
                     "
     primer_bind     complement(346..386)
                     /label=Human_Dlx_R_Gibson
     misc_feature    346..370
                     /label=Gibson_hDlx_I_R
     misc_feature    370
                     /label=h cloning
     misc_feature    371..704
                     /label=m56ii_enhancer
     misc_feature    371..386
                     /label=Gibson_hDlx_P_R
     misc_feature    371
                     /label=""C"" after PpumI digest
     primer_bind     514..533
                     /label=Seq_AAV_hDlx_F2
     primer_bind     complement(514..533)
                     /label=AAV_hDlx_R2
     promoter        740..788
                     /label=pB-Globin
     primer_bind     complement(743..762)
                     /label=AAV_hDlx_R1
     misc_feature    835..1005
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     intron          873..1005
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=Seq_AAV_hDlx_F3
     misc_feature    1006..1047
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     primer_bind     1012..1031
                     /label=Seq_AAV_Dlx_F
     misc_feature    1023..1042
                     /label=Gibson_NLS_F1a
     misc_feature    1026..1041
                     /label=Gibson_F_16bp
     protein_bind    1048..1081
                     /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    1082..1253
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    1082..1095
                     /label=Gibs_Flex_Fb
     protein_bind    1088..1121
                     /label=Lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites."
     protein_bind    1172..1205
                     /label=LoxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    1194..1213
                     /label=Gibson_Flex_R1
     misc_feature    1214..1219
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    1214..1219
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     misc_feature    1214..1219
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    1214..1219
                     /label=SpeI
     misc_feature    1220..1253
                     /label=MCS
                     /note=" multiple cloning site
                     "
     primer_bind     1238..1282
                     /label=GA-GFP.Rev
     misc_feature    1259..1984
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     CDS             complement(1259..1978)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=eGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     1917..1933
                     /label=pEGFP_N_Macrogen_Seq
     misc_feature    1957..1984
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     primer_bind     complement(1966..2004)
                     /label=GA-GFP.For
     misc_feature    1979..1984
                     /label=Kozak
     misc_feature    1990..2145
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    1990..2008
                     /label=MCS
                     /note=" multiple cloning site
                     "
     misc_feature    2009..2015
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     misc_feature    2009..2015
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    2010..2015
                     /label=SpeI
     misc_feature    2028..2061
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     protein_bind    complement(2028..2061)
                     /label=Lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites."
     misc_feature    2054..2061
                     /label=Gibs_Flex_Fb
     misc_feature    2112..2145
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     protein_bind    complement(2112..2145)
                     /label=LoxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    2112..2123
                     /label=Gibson_Flex_R1
     protein_bind    2146..2179
                     /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    2188..2776
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     polyA_signal    2784..3008
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3010..3150
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      3225..3680
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        3962..4066
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4067..4927
                     /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      5098..5686
                     /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 agagggcccg cggccgcacg cgtttattca gaatgttatg
      181 cactcacagt ggtttggcat gcatctggtg aatttttttt aacgaaaaat tagtgttggt
      241 ttcgatgtat ggtagcattc tccctaacgt aatttgaata attcagcaaa gccccactac
      301 cagctgtact tctgcagcct cttccattct tttcagcatt ataattttgg ttaattttca
      361 attttaggtc ctacgtctct gcaatttgtg tatgaataac agaataattt ccctcttttg
      421 tttcgccttt cctgttcctg aatctaaata aagatggctt tttagtatta aaagtggaag
      481 aaaattacag gtaattatct ttgacggtaa aaacgctgta atcagcgggc tacatgaaaa
      541 attactctaa ttatggctgc atttaagaga atggaaaaaa accttcttgt ggataaaaac
      601 cttaaattgt ccccaatgtc tgcttcaaat tggatggcac tgcagctgga ggctttgttc
      661 agaattgatc ctggggagct acgaacccaa agtttcacag taggaagcta gatatcgttt
      721 aaacttcctg cagcccgggc tgggcataaa agtcagggca gagccatcta ttgcttacat
      781 ttgcttctag cctgcaagtc gaggagcgca gccttccaga agcagagcgc ggcgccttaa
      841 gctgcagaag ttggtcgtga ggcactgggc acgtaagtat caaggttaca agacaggttt
      901 aaggagacca atagaaactg ggcttgtcga gacagagaag actcttgcgt ttctgatagg
      961 cacctattgg tcttactgac atccactttg cctttctctc cacaggtgtc cactcccagt
     1021 tcaattacag ctcttaagaa agctagcgaa gttcctattc tctagaaagt ataggaactt
     1081 ctccggaata acttcgtata ggatacttta tacgaagtta tgcagaatgg tagctggatt
     1141 gtagctgcta ttagcaatat gaaacctctt aataacttcg tatagcatac attatacgaa
     1201 gttataacgc gccactagtg aattcgagct cggtacccgg ggatcctcta gagtcgactt
     1261 acttgtacag ctcgtccatg ccgagagtga tcccggcggc ggtcacgaac tccagcagga
     1321 ccatgtgatc gcgcttctcg ttggggtctt tgctcagggc ggactgggtg ctcaggtagt
     1381 ggttgtcggg cagcagcacg gggccgtcgc cgatgggggt gttctgctgg tagtggtcgg
     1441 cgagctgcac gctgccgtcc tcgatgttgt ggcggatctt gaagttcacc ttgatgccgt
     1501 tcttctgctt gtcggccatg atatagacgt tgtggctgtt gtagttgtac tccagcttgt
     1561 gccccaggat gttgccgtcc tccttgaagt cgatgccctt cagctcgatg cggttcacca
     1621 gggtgtcgcc ctcgaacttc acctcggcgc gggtcttgta gttgccgtcg tccttgaaga
     1681 agatggtgcg ctcctggacg tagccttcgg gcatggcgga cttgaagaag tcgtgctgct
     1741 tcatgtggtc ggggtagcgg ctgaagcact gcacgccgta ggtcagggtg gtcacgaggg
     1801 tgggccaggg cacgggcagc ttgccggtgg tgcagatgaa cttcagggtc agcttgccgt
     1861 aggtggcatc gccctcgccc tcgccggaca cgctgaactt gtggccgttt acgtcgccgt
     1921 ccagctcgac caggatgggc accaccccgg tgaacagctc ctcgcccttg ctcaccatgg
     1981 tggcgtcgac ctgcaggcat gcaagcttta ctagtggtgg cgctagcata acttcgtata
     2041 aagtatccta tacgaagtta tttgccttaa cccagaaatt atcactgtta ttctttagaa
     2101 tggtgcaaag aataacttcg tataatgtat gctatacgaa gttatgaagt tcctattctc
     2161 tagaaagtat aggaacttcg gcgcgccaat caacctctgg attacaaaat ttgtgaaaga
     2221 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     2281 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     2341 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     2401 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     2461 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     2521 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     2581 aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     2641 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     2701 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     2761 tgggccgcct ccccgcctgg atcctgtgcc ttctagttgc cagccatctg ttgtttgccc
     2821 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     2881 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
     2941 gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg
     3001 ctctatggca ggaaccccta gtgatggagt tggccactcc ctctctgcgc gctcgctcgc
     3061 tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag
     3121 tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc
     3181 atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg
     3241 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3301 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3361 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3421 cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3481 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3541 tggaacaaca ctcaacccta tctcgggcta ttcttttgat ttataaggga ttttgccgat
     3601 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     3661 aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg atgccgcata
     3721 gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     3781 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     3841 ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata
     3901 ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt
     3961 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     4021 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     4081 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     4141 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     4201 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     4261 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     4321 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     4381 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     4441 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     4501 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     4561 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     4621 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     4681 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     4741 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     4801 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     4861 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     4921 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     4981 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     5041 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5101 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5161 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5221 cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa
     5281 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5341 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     5401 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     5461 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     5521 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     5581 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     5641 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     5701 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgt
//
