LOCUS       Exported                6515 bp ds-DNA     circular SYN 16-MAR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6515)
  AUTHORS   SnapGene License
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Mar 20, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6515
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          1493..3228
                     /organism="Homo sapiens"
                     /chromosome="8"
                     /map="8p23.3"
                     /mol_type="mRNA"
                     /db_xref="taxon:9606"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     primer_bind     141..190
                     /label=F-D1
     primer_bind     147..194
                     /label=F-Mlu-D2
     primer_bind     147..161
                     /label=3'-pcalm-ampF
     primer_bind     150..168
                     /label=P-Calm1-F
     primer_bind     151..200
                     /label=F-D2
     misc_feature    156..276
                     /label=P Calm1
     misc_feature    156..165
                     /label=D1
     primer_bind     161..210
                     /label=F-D3
     primer_bind     complement(161..185)
                     /label=R-D4
     misc_feature    166..175
                     /label=D2
     primer_bind     171..220
                     /label=F-D4
     primer_bind     complement(171..195)
                     /label=R-D5
     misc_feature    176..185
                     /label=D3
     primer_bind     181..230
                     /label=F-D5
     primer_bind     complement(181..205)
                     /label=R-D6
     misc_feature    186..195
                     /label=D4
     primer_bind     191..240
                     /label=F-D6
     primer_bind     complement(191..215)
                     /label=r-d7
     misc_feature    196..205
                     /label=D5
     primer_bind     200..250
                     /label=F-D7
     primer_bind     complement(201..225)
                     /label=R-D8
     misc_feature    206..215
                     /label=D6
     primer_bind     211..260
                     /label=F-D8
     primer_bind     complement(215..236)
                     /label=R-D9
     misc_feature    216..225
                     /label=D7
     primer_bind     222..271
                     /label=F-D9
     primer_bind     complement(222..246)
                     /label=R-D10
     misc_feature    226..235
                     /label=D8
     primer_bind     complement(232..256)
                     /label=R-D11
     misc_feature    236
                     /label=+1
     misc_feature    237..246
                     /label=D9
     primer_bind     complement(242..266)
                     /label=R-D12
     misc_feature    247..256
                     /label=D10
     misc_feature    257..266
                     /label=D11
     primer_bind     complement(260..287)
                     /label=2B-1R
     primer_bind     complement(261..276)
                     /label=P-Calm1-R
     misc_feature    267..276
                     /label=D12
     primer_bind     271..296
                     /label=2B-2F
     primer_bind     complement(277..298)
                     /label=Primer 1
     misc_feature    277..282
                     /label=BsiWI
     primer_bind     283..302
                     /label=Primer 5
     CDS             288..1318
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     misc_feature    548
                     /label=silent mut
     primer_bind     complement(616..650)
                     /label=R-Cre-R119STOP
     primer_bind     complement(632..651)
                     /label=R-CRE-L115-BamHI
     primer_bind     636..670
                     /label=F-Cre-R119STOP
     primer_bind     1093..1116
                     /label=F-Seq-Cre-3F1
     misc_feature    1093..1098
                     /label=ClaI
     misc_feature    1224
                     /label=mut
     primer_bind     1262..1279
                     /label=cre-SF
     primer_bind     complement(1296..1318)
                     /label=R-T5-Cre
     intron          1320..1360
                     /label=Splicing donor
                     /note="Splicing donor"
     primer_bind     complement(1338..1367)
                     /label=2B-2R
     primer_bind     complement(1341..1385)
                     /label=Primer 7
     primer_bind     complement(1341..1385)
                     /label=Primer 6
     primer_bind     1350..1387
                     /label=2B-3F
     primer_bind     complement(1356..1380)
                     /label=Primer 3
     primer_bind     1367..1401
                     /label=Primer 8
     primer_bind     1367..1401
                     /label=Primer 2
     primer_bind     1367..1401
                     /label=F-T5-JT+2272
     protein_bind    1367..1400
                     /label=loxJTZ17?2272?
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    1380..1387
                     /label=2272
     intron          1401..1492
                     /label=Splicing acceptor
                     /note="Splicing acceptor"
     primer_bind     1401..1420
                     /label=Primer 4
     primer_bind     1459..1478
                     /label=T0060_PIRES-F
     primer_bind     complement(1468..1492)
                     /label=#57-R-SA-3R1
     primer_bind     1477..1515
                     /label=B9F-NheI-F
     primer_bind     1477..1492
                     /label=Primer 10
     misc_feature    1492
                     /label=NheI?
     CDS             1493..3228
                     /codon_start=1
                     /gene="ARHGEF10"
                     /gene_synonym="GEF10; SNCV"
                     /product="rho guanine nucleotide exchange factor 10
                     isoform 1"
                     /label=ARHGEF10
                     /note="isoform 1 is encoded by transcript variant 1; Rho
                     guanine nucleotide exchange factor (GEF) 10"
                     /db_xref="GI:62548864"
                     /db_xref="CCDS:CCDS34794.1"
                     /db_xref="GeneID:9639"
                     /db_xref="HGNC:HGNC:14103"
                     /db_xref="MIM:608136"
                     /protein_id="NP_055444.2"
                     /translation="LQRLILKKEDEIRAADCCRIQLQLPGKQDKSGRPTFFTAVFNTFT
                     PAIKESWVNSLQMAKLALEEENHMGWFCVEDDGNHIKKEKHPLLVGHMPVMVAKQQEFK
                     IECAAYNPEPYLNNESQPDSFSTAHGFLWIGSCTHQMGQIAIVSFQNSTPKVIECFNVE
                     SRILCMLYVPVEEKRREPGAPPDPETPAVRASDVPTICVGTEEGSISIYKSSQGSKKVR
                     LQHFFTPEKSTVMSLACTSQSLYAGLVNGAVASYARAPDGSWDSEPQKVIKLGVLPVRS
                     LLMMEDTLWAASGGQVFIISVETHAVEGQLEAHQEEGMVISHMAVSGVGIWIAFTSGST
                     LRLFHTETLKHLQDINIATPVHNMLPGHQRLSVTSLLVCHGLLMVGTSLGVLVALPVPR
                     LQGIPKVTGRGMVSYHAHNSPVKFIVLATALHEKDKDKSRDSLAPGPEPQDEDQKDALP
                     SGGAGSSLSQGDPDAAIWLGDSLGSMTQKSDLSSSSGSLSLSHGSSSLEHRSEDSTIYD
                     LLKDPVSLRSKARRAKKAKASSALVVCGGQGHRRVHRKARQPHQEELAPTVMVWQIPLL
                     NI"
     misc_feature    1493..1494
                     /label=Feature 35
     primer_bind     complement(2530..2546)
                     /label=B9F-SR
     primer_bind     2779..2790
                     /label=P-Calm1-F
     primer_bind     complement(3209..3249)
                     /label=B9F-BsrGI-R
     misc_feature    3229..3234
                     /label=BsrGI
     CDS             3235..3300
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3240..3328)
                     /label=296
     terminator      3304..3735
                     /label=W3SL
                     /note="Short WPRE and SV40 polyA"
     primer_bind     3304..3329
                     /label=#56-F-W3SL-5F1
     primer_bind     complement(3383..3406)
                     /label=R-Seq-W3SL-5R1
     misc_feature    3778..3918
                     /label=R-ITR
     rep_origin      3921..4376
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4068..4077)
                     /label=F-Seq-Cre-3F1
     promoter        4658..4762
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4763..5623
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="HSNMYPLMRQ*P**MLQ*Y*KRKSMSIQHFRVALIPFFAAFCLPV
                     FAHPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLS
                     RIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTT
                     IGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLT
                     LASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIV
                     VIYTTG"
     rep_origin      5794..6382
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtcgctc tgcgcgaggc ggctccgccg
      181 cggcggaggg atacggcgca ccatatatat atcgcggggc gcagactcgc gctccggcag
      241 tggtgctggg agtgtcgtgg acgccgtgcc gttactcgta cgccaccatg tccaatttac
      301 tgaccgtaca ccaaaatttg cctgcattac cggtcgatgc aacgagtgat gaggttcgca
      361 agaacctgat ggacatgttc agggatcgcc aggcgttttc tgagcatacc tggaaaatgc
      421 ttctgtccgt ttgccggtcg tgggcggcat ggtgcaagtt gaataaccgg aaatggtttc
      481 ccgcagaacc tgaagatgtt cgcgattatc ttctatatct tcaggcgcgc ggtctggcag
      541 taaaaaccat ccagcaacat ttgggccagc taaacatgct tcatcgtcgg tccgggctgc
      601 cacgaccaag tgacagcaat gctgtttcac tggttatgcg gcggatccga aaagaaaacg
      661 ttgatgccgg tgaacgtgca aaacaggctc tagcgttcga acgcactgat ttcgaccagg
      721 ttcgttcact catggaaaat agcgatcgct gccaggatat acgtaatctg gcatttctgg
      781 ggattgctta taacaccctg ttacgtatag ccgaaattgc caggatcagg gttaaagata
      841 tctcacgtac tgacggtggg agaatgttaa tccatattgg cagaacgaaa acgctggtta
      901 gcaccgcagg tgtagagaag gcacttagcc tgggggtaac taaactggtc gagcgatgga
      961 tttccgtctc tggtgtagct gatgatccga ataactacct gttttgccgg gtcagaaaaa
     1021 atggtgttgc cgcgccatct gccaccagcc agctatcaac tcgcgccctg gaagggattt
     1081 ttgaagcaac tcatcgattg atttacggcg ctaaggatga ctctggtcag agatacctgg
     1141 cctggtctgg acacagtgcc cgtgtcggag ccgcgcgaga tatggcccgc gctggagttt
     1201 caataccgga gatcatgcaa gctggtggct ggaccaatgt aaatattgta atgaactata
     1261 tccgtaacct ggatagtgaa acaggggcaa tggtgcgcct gctggaagat ggcgattagg
     1321 taagtatcaa ggttacaaga caggtttaag gagaccaata actagtataa cttcgtatag
     1381 gatactttat agcaatttat gaaactgggc ttgtcgagac agagaagact cttgcgtttc
     1441 tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac aggtttacaa
     1501 aggcttattt tgaagaaaga agatgaaatc agagctgcgg actgctgcag aattcagtta
     1561 cagcttcccg ggaagcagga caaatctggg cgaccgacgt tctttacagc tgtgttcaat
     1621 acgttcactc ctgccatcaa ggagtcctgg gtcaacagct tacagatggc caagctcgcc
     1681 ctagaagagg agaaccacat gggctggttc tgtgtggaag acgatgggaa tcacattaaa
     1741 aaggagaagc atcctctcct cgtcggacac atgcccgtga tggtggccaa gcagcaggag
     1801 ttcaagattg aatgtgctgc ttataaccct gaaccttacc taaataatga aagccagcca
     1861 gattcatttt ccacggcaca tggtttcctg tggatcggaa gttgcaccca tcaaatgggt
     1921 cagattgcca tcgtctcgtt tcaaaattcc actcccaaag tcattgagtg cttcaacgtg
     1981 gaatctcgca tcctgtgcat gctgtacgtt cccgtcgagg agaagcgcag agagcctggg
     2041 gcacccccgg accccgagac cccggccgtg agagcttctg atgtccccac gatctgtgta
     2101 gggacggagg agggaagcat ttccatttat aaaagcagtc aaggctccaa gaaagtgaga
     2161 cttcagcact ttttcactcc tgagaagtcc acagtcatga gcctggcttg cacgtctcag
     2221 agcctgtacg ctggcctggt caacggggca gtcgccagct acgccagagc cccagatgga
     2281 tcctgggatt cagaacctca aaaagtgatc aagttaggcg tcctaccagt tagaagtcta
     2341 ctcatgatgg aagacacgtt gtgggcggct tccggaggtc aagtcttcat catcagtgtg
     2401 gagactcatg ctgtagaggg tcagctggag gcccaccagg aggaaggcat ggtgatctcc
     2461 cacatggccg tgtccggcgt cgggatctgg attgccttca cctcagggtc cacgctccgc
     2521 ctttttcaca cggaaactct caagcacctg caggacatca acatcgccac ccctgttcac
     2581 aacatgctgc cagggcacca gcggctgtcg gtgacgagcc tgctcgtctg ccacggattg
     2641 ctgatggtcg gcaccagcct gggagtcctc gtggccctgc cggtcccacg tctgcaaggg
     2701 attcccaaag tgaccggaag aggcatggtc tcctaccatg cacacaacag tcctgtcaaa
     2761 ttcatcgtcc tggccacggc tctgcacgag aaagacaagg acaaatccag ggacagcctg
     2821 gctcctggcc ccgagcctca ggacgaagac cagaaggacg cacttccgag tggaggagct
     2881 ggttcatctc tgagccaggg tgaccctgac gcagccatct ggttgggaga ttcgctggga
     2941 tcgatgactc agaaaagcga cctgtcctcc tcatctgggt ccctgagctt gtctcacggc
     3001 tccagctctc tagagcacag atcagaggac agcaccatct atgatctcct gaaggatcct
     3061 gtctcgctga gaagcaaagc acgccgggcc aagaaagcca aggccagctc ggcgctggtg
     3121 gtctgtggag ggcagggcca ccgccgggtg cacaggaagg cccggcagcc ccaccaggaa
     3181 gagctggcgc cgaccgtcat ggtctggcag atccctctgc tgaatatatg tacagactac
     3241 aaagaccatg acggtgatta taaagatcat gacatcgact acaaggatga cgatgacaag
     3301 taactcgaga taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     3361 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     3421 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggtta gttcttgcca
     3481 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     3541 ctgacaattc cgtggtgttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     3601 tctagcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc
     3661 aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggagatg
     3721 tgggaggttt tttaaagatc tggagaccgg tgtggcacgt gcggaccgag cggccgcagg
     3781 aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg
     3841 ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag
     3901 cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     3961 tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc
     4021 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc
     4081 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4141 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4201 acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     4261 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     4321 caaccctatc tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg
     4381 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt
     4441 tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     4501 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     4561 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     4621 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     4681 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     4741 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     4801 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     4861 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     4921 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     4981 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     5041 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     5101 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     5161 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     5221 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     5281 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     5341 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     5401 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     5461 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     5521 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     5581 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     5641 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     5701 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     5761 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     5821 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5881 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     5941 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     6001 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6061 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6121 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6181 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6241 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6301 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     6361 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     6421 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     6481 gttcctggcc ttttgctggc cttttgctca catgt
//
