LOCUS       Exported                7617 bp DNA     circular SYN 14-DEC-2023
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 7617)
  AUTHORS   Fishell Lab
  TITLE     Direct Submission
  JOURNAL   Exported Dec 14, 2023 from SnapGene 6.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7617
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     repeat_region   1..130
                     /label=L-ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     misc_feature    108..1282
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     misc_feature    132..144
                     /label=Gibson_hDlx_P_F
     misc_feature    139
                     /label=A from MluI
     misc_feature    140..150
                     /label=Reconstituted MluI
     misc_feature    145..150
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     enhancer        151..987
                     /label=S9E27
     primer_bind     complement(694..714)
                     /label=S9E27_R_seq_JM
     misc_feature    988..997
                     /label=Reconstituted PmeI
     misc_feature    988..993
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_G_EGFP-N"
     promoter        1017..1065
                     /label=pB-Globin
     misc_feature    1112..1282
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     intron          1150..1282
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1290..1351
                     /label=Gibson_HA-NLS_F1
     misc_feature    1291..3929
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    1296..1301
                     /label=Kozak
     CDS             1302..3098
                     /codon_start=1
                     /product="From Roth LabHA tag added
                     "
                     /label=HA-GqDREADD
                     /translation="MYPYDVPDYATLHNNSTTSPLFPNISSSWIHSPSDAGLPPGTVTH
                     FGSYNVSRAAGNFSSPDGTTDDPLGGHTVWQVVFIAFLTGILALVTIIGNILVIVSFKV
                     NKQLKTVNNYFLLSLACADLIIGVISMNLFTTYIIMNRWALGNLACDLWLAIDCVASNA
                     SVMNLLVISFDRYFSITRPLTYRAKRTTKRAGVMIGLAWVISFVLWAPAILFWQYFVGK
                     RTVPPGECFIQFLSEPTITFGTAIAGFYMPVTIMTILYWRIYKETEKRTKELAGLQASG
                     TEAETENFVHPTGSSRSCSSYELQQQSMKRSNRRKYGRCHFWFTTKSWKPSSEQMDQDH
                     SSSDSWNNNDAAASLENSASSDEEDIGSETRAIYSIVLKLPGHSTILNSTKLPSSDNLQ
                     VPEEELGMVDLERKADKLQAQKSVDDGGSFPKSFSKLPIQLESAVDTAKTSDVNSSVGK
                     STATLPLSFKEATLAKRFALKTRSQITKRKRMSLVKEKKAAQTLSAILLAFIITWTPYN
                     IMVLVNTFCDSCIPKTFWNLGYWLCYINSTVNPVCYALCNKTFRTTFKMLLLCQCDKKK
                     RRKQQYQQRQSVIFHKRAPEQAL"
     misc_feature    1305..1331
                     /label=140708_Sequencing_Clone1
                     /note="Check 140708_AAV_GFP_1_R_140619_Seq_AAV_Dlx_R"
     misc_feature    1332..1351
                     /label=Gibson_NLS_F1b
     primer_bind     1711..1730
                     /label=Seq_AAV_hDlx_F4
     primer_bind     2240..2259
                     /label=Seq_AAV_hDlx_F5
     misc_feature    2369
                     
     misc_feature    2417
                     /label=Missing in Clone 3 F
                     /note="Could be double peak on Raw"
     misc_feature    2498
                     /label=End of sequencing Clone1
     misc_feature    2693
                     /label=End of sequencing Clone 1 R
     primer_bind     2832..2851
                     /label=Seq_AAV_hDlx_F6
     primer_bind     3040..3059
                     /label=Seq_AAV_hDlx_F7
     primer_bind     complement(3079..3124)
                     /label=Gibson_NLS_R1
     primer_bind     3079..3124
                     /label=Gibson_NLS_F2
     misc_feature    3079..3098
                     /label=Gibson_NLS_R1
     misc_feature    3086..3098
                     /label=Overlap Gibson 2A
     misc_feature    3096..3098
                     /label=150212_AAV_hDLX_F7_Sequenced
                     /note="Ok for the 12 minis"
     misc_feature    3099..3104
                     /label=BspEI
     misc_feature    3105..3124
                     /label=Gibson_NLS_F2
     CDS             3111..3176
                     /codon_start=1
                     /label=P2A
                     /translation="GSGATNFSLLKQAGDVEENPGP"
     CDS             3189..3209
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3210..3908
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGL
                     VTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIHQ
                     ALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRHH
                     LFLYGMDELYK"
     primer_bind     complement(3251..3270)
                     /label=Seq-mCherry5-R
     primer_bind     complement(3892..3944)
                     /label=140620_eGFP_R_Gibson
     misc_feature    3904..3923
                     /label=Gibson_NLS_R2
     misc_feature    3924..3943
                     /label=Gibson_NLS_R2b
     misc_feature    3926..3941
                     /label=Gibson_R_16bp
     misc_feature    3927
                     /label=begening of sequencing CLone1 with R
     misc_feature    3963..4551
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     polyA_signal    4720..4801
                     /label=SV40pA
                     /note="SV40 polyadenylation signal"
     repeat_region   4880..5020
                     /label=R-ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      5095..5550
                     /direction=RIGHT
                     /label=F1 Origin
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        5832..5936
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5937..6797
                     /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      6968..7556
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7574..7593
                     /label=Seq_AAV_hDlx_F1
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtatcgat cctcgggtaa tttgtaaaga caatttttct
      181 tttccccctc tgaacctagg ccttaggctg gctaggcaag tctccactac agccccgccc
      241 actccatctc ggctgtggct cgcgggaaag gcagccccct acagttagtc tccccagcca
      301 tcttctccgg ttcccctgat gctctcatgg ccccagccat cctagagctg gtgcgccatc
      361 gctcaccttt agcattcggc ctgctggggg tgagcttttg ggttagaagg aacaaggcat
      421 cagatggggt cgtgtttttt tacattccaa tcagttttcg gaggggaaga acagacattt
      481 ttgtcagcta attgcattta attacagaga caatcgctgt ttccgaacgt tgttgtgtcc
      541 actccctcat ccttcaatca gttcaagcac atccggcccc agctgggagc cttttaattg
      601 cataatttgg atttcacttc tggaaagaaa cttgagaaag actccgtgag ggtctccatt
      661 aacacaatca gctccccagg cacccggggc tggccctaat tgacccatct cagctaatgc
      721 ctctttccaa agaacgcccg cgtcatttct ttggcttgtt tgtgctattg tatcctgttt
      781 tagtgtcttg atcttaaaca tctttaaaga gggcactcag tggcttgttt aactggaagt
      841 ggtccagctg gtttggaagg acagtgcatc aggtgatgta agtccaaggg taggaatttc
      901 tggcctctcc ttcccctgcc cattgcaccg ctctccccaa caggaagtag ctacacatga
      961 taaggtgcac tgtggcttct aggacttatc gatgtttaaa cttcctgcag cccgggctgg
     1021 gcataaaagt cagggcagag ccatctattg cttacatttg cttctagcct gcaagtcgag
     1081 gagcgcagcc ttccagaagc agagcgcggc gccttaagct gcagaagttg gtcgtgaggc
     1141 actgggcacg taagtatcaa ggttacaaga caggtttaag gagaccaata gaaactgggc
     1201 ttgtcgagac agagaagact cttgcgtttc tgataggcac ctattggtct tactgacatc
     1261 cactttgcct ttctctccac agcgagctcg ctagcgccac catgtaccca tacgatgttc
     1321 cagattacgc taccttgcac aataacagta caacctcgcc tttgtttcca aacatcagct
     1381 cctcctggat acacagcccc tccgatgcag ggctgccccc gggaaccgtc actcatttcg
     1441 gcagctacaa tgtttctcga gcagctggca atttctcctc tccagacggt accaccgatg
     1501 accctctggg aggtcatacc gtctggcaag tggtcttcat cgctttctta acgggcatcc
     1561 tggccttggt gaccatcatc ggcaacatcc tggtaattgt gtcatttaag gtcaacaagc
     1621 agctgaagac ggtcaacaac tacttcctct taagcctggc ctgtgccgat ctgattatcg
     1681 gggtcatttc aatgaatctg tttacgacct acatcatcat gaatcgatgg gccttaggga
     1741 acttggcctg tgacctctgg cttgccattg actgcgtagc cagcaatgcc tctgttatga
     1801 atcttctggt catcagcttt gacagatact tttccatcac gaggccgctc acgtaccgag
     1861 ccaaacgaac aacaaagaga gccggtgtga tgatcggtct ggcttgggtc atctcctttg
     1921 tcctttgggc tcctgccatc ttgttctggc aatactttgt tggaaagaga actgtgcctc
     1981 cgggagagtg cttcattcag ttcctcagtg agcccaccat tacttttggc acagccatcg
     2041 ctggttttta tatgcctgtc accattatga ctattttata ctggaggatc tataaggaaa
     2101 ctgaaaagcg taccaaagag cttgctggcc tgcaagcctc tgggacagag gcagagacag
     2161 aaaactttgt ccaccccacg ggcagttctc gaagctgcag cagttacgaa cttcaacagc
     2221 aaagcatgaa acgctccaac aggaggaagt atggccgctg ccacttctgg ttcacaacca
     2281 agagctggaa acccagctcc gagcagatgg accaagacca cagcagcagt gacagttgga
     2341 acaacaatga tgctgctgcc tccctggaga actccgcctc ctccgacgag gaggacattg
     2401 gctccgagac gagagccatc tactccatcg tgctcaagct tccgggtcac agcaccatcc
     2461 tcaactccac caagttaccc tcatcggaca acctgcaggt gcctgaggag gagctgggga
     2521 tggtggactt ggagaggaaa gccgacaagc tgcaggccca gaagagcgtg gacgatggag
     2581 gcagttttcc aaaaagcttc tccaagcttc ccatccagct agagtcagcc gtggacacag
     2641 ctaagacttc tgacgtcaac tcctcagtgg gtaagagcac ggccactcta cctctgtcct
     2701 tcaaggaagc cactctggcc aagaggtttg ctctgaagac cagaagtcag atcactaagc
     2761 ggaaaaggat gtccctggtc aaggagaaga aagcggccca gaccctcagt gcgatcttgc
     2821 ttgccttcat catcacttgg accccataca acatcatggt tctggtgaac accttttgtg
     2881 acagctgcat acccaaaacc ttttggaatc tgggctactg gctgtgctac atcaacagca
     2941 ccgtgaaccc cgtgtgctat gctctgtgca acaaaacatt cagaaccact ttcaagatgc
     3001 tgctgctgtg ccagtgtgac aaaaaaaaga ggcgcaagca gcagtaccag cagagacagt
     3061 cggtcatttt tcacaagcgc gcacccgagc aggccttgtc cggagataac ggaagcggag
     3121 ccactaactt ctccctgttg aaacaagcag gggatgtcga agagaatccc gggccatcta
     3181 gaatggcccc caagaagaag aggaaggtgg tgagcaaggg cgaggaggtc atcaaagagt
     3241 tcatgcgctt caaggtgcgc atggagggct ccatgaacgg ccacgagttc gagatcgagg
     3301 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     3361 gcggccccct gcccttcgcc tgggacatcc tgtcccccca gttcatgtac ggctccaagg
     3421 cgtacgtgaa gcaccccgcc gacatccccg attacaagaa gctgtccttc cccgagggct
     3481 tcaagtggga gcgcgtgatg aacttcgagg acggcggtct ggtgaccgtg acccaggact
     3541 cctccctgca ggacggcacg ctgatctaca aggtgaagat gcgcggcacc aacttccccc
     3601 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctccacc gagcgcctgt
     3661 acccccgcga cggcgtgctg aagggcgaga tccaccaggc cctgaagctg aaggacggcg
     3721 gccactacct ggtggagttc aagaccatct acatggccaa gaagcccgtg caactgcccg
     3781 gctactacta cgtggacacc aagctggaca tcacctccca caacgaggac tacaccatcg
     3841 tggaacagta cgagcgctcc gagggccgcc accacctgtt cctgtacggc atggacgagc
     3901 tgtacaagta agtcgacggc gcgccccacc cctgcaggga attcgatatc aagcttatcg
     3961 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     4021 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     4081 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     4141 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     4201 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     4261 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     4321 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     4381 tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     4441 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     4501 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     4561 gagcgctgct cgagagatct gtgatagcgg ccatcaagct ggccgcgact ctagatcata
     4621 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     4681 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     4741 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4801 cattctagtt gtggtttgtc caaactcatc aatgtatcag cttatcgata ccgcatgcac
     4861 gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc ctctctgcgc
     4921 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg
     4981 gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt
     5041 ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc
     5101 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     5161 ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     5221 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     5281 tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc
     5341 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     5401 tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat ttataaggga
     5461 ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     5521 attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg
     5581 atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     5641 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     5701 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
     5761 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     5821 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     5881 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     5941 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     6001 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     6061 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     6121 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     6181 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     6241 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     6301 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     6361 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     6421 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     6481 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     6541 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6601 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6661 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6721 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6781 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     6841 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     6901 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     6961 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     7021 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     7081 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     7141 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     7201 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     7261 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     7321 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     7381 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     7441 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     7501 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     7561 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgt
//
