LOCUS       Exported                6465 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 6465)
  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..6465
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          1493..3178
                     /organism="Homo sapiens"
                     /chromosome="21"
                     /map="21q22.11"
                     /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..1513
                     /label=B48F-NheI-F
     primer_bind     1477..1492
                     /label=Primer 10
     misc_feature    1492
                     /label=NheI?
     CDS             1493..3178
                     /codon_start=1
                     /gene="ITSN1"
                     /gene_synonym="ITSN; SH3D1A; SH3P17"
                     /product="intersectin-1 isoform ITSN-l"
                     /label=ITSN1
                     /note="isoform ITSN-l is encoded by transcript variant 1;
                     human intersectin-SH3 domain-containing protein SH3P17; Src
                     homology 3 domain-containing protein; SH3 domain-containing
                     protein 1A; intersectin 1 (SH3 domain protein)"
                     /db_xref="GI:47717123"
                     /db_xref="CCDS:CCDS33545.1"
                     /db_xref="GeneID:6453"
                     /db_xref="HGNC:HGNC:6183"
                     /db_xref="MIM:602442"
                     /protein_id="NP_003015.2"
                     /translation="VIGMYDYTAQNDDELAFNKGQIINVLNKEDPDWWKGEVNGQVGLF
                     PSNYVKLTTDMDPSQQWCSDLHLLDMLTPTERKRQGYIHELIVTEENYVNDLQLVTEIF
                     QKPLMESELLTEKEVAMIFVNWKELIMCNIKLLKALRVRKKMSGEKMPVKMIGDILSAQ
                     LPHMQPYIRFCSRQLNGAALIQQKTDEAPDFKEFVKRLAMDPRCKGMPLSSFILKPMQR
                     VTRYPLIIKNILENTPENHPDHSHLKHALEKAEELCSQVNEGVREKENSDRLEWIQAHV
                     QCEGLSEQLVFNSVTNCLGPRKFLHSGKLYKAKSNKELYGFLFNDFLLLTQITKPLGSS
                     GTDKVFSPKSNLQYKMYKTPIFLNEVLVKLPTDPSGDEPIFHISHIDRVYTLRAESINE
                     RTAWVQKIKAASELYIETEKKKREKAYLVRSQRATGIGRLMVNVVEGIELKPCRSHGKS
                     NPYCEVTMGSQCHITKTIQDTLNPKWNSNCQFFIRDLEQEVLCITVFERDQFSPDDFLG
                     RTEIRVADIKKDQGSKGPVTKCLLLHEVPTGEIVVRLDLQLFDEP"
     misc_feature    1493..1494
                     /label=Feature 66
     primer_bind     2092..2108
                     /label=B48F-SF
     primer_bind     complement(3160..3199)
                     /label=B48F-BsrGI-R
     misc_feature    3179..3184
                     /label=BsrGI
     CDS             3185..3250
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3190..3278)
                     /label=296
     terminator      3254..3685
                     /label=W3SL
                     /note="Short WPRE and SV40 polyA"
     primer_bind     3254..3279
                     /label=#56-F-W3SL-5F1
     primer_bind     complement(3333..3356)
                     /label=R-Seq-W3SL-5R1
     misc_feature    3728..3868
                     /label=R-ITR
     rep_origin      3871..4326
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4018..4027)
                     /label=F-Seq-Cre-3F1
     promoter        4608..4712
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4713..5573
                     /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      5744..6332
                     /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 aggtgattgg
     1501 gatgtacgac tacaccgcgc agaatgacga tgagctggcc ttcaacaagg gccagatcat
     1561 caacgtcctc aacaaggagg accctgactg gtggaaagga gaagtcaatg gacaagtggg
     1621 gctcttccca tccaattatg tgaagctgac cacagacatg gacccaagcc agcaatggtg
     1681 ttcagactta catctcttgg atatgttgac cccaactgaa agaaagcgac aaggatacat
     1741 ccacgagctc attgtcaccg aggagaacta tgtgaatgac ctgcagctgg tcacagagat
     1801 ttttcaaaaa cccctgatgg agtctgagct gctgacagaa aaagaggttg ctatgatttt
     1861 tgtgaactgg aaggagctga ttatgtgtaa tatcaaacta ctaaaagcgc tgagagtccg
     1921 caagaagatg tccggggaga agatgcctgt gaagatgatt ggagacatcc tgagcgcaca
     1981 gctgccgcac atgcagccct acatccgctt ctgcagccgc cagctcaacg gggctgccct
     2041 gatccagcag aagacggacg aggccccaga cttcaaggag ttcgtcaaaa gattggcaat
     2101 ggatcctcgg tgtaaaggga tgccactctc tagttttata ctgaagccta tgcaacgggt
     2161 aacaagatac ccactgatca ttaaaaatat cctggaaaac acccctgaaa accacccgga
     2221 ccacagccac ttgaagcacg ccctggagaa ggcggaagag ctctgttccc aggtgaacga
     2281 aggggtgcgg gagaaggaga actctgaccg gctggagtgg atccaggccc acgtgcagtg
     2341 tgaaggcctg tctgagcaac ttgtgttcaa ttcagtgacc aattgcttgg ggccgcgcaa
     2401 atttctgcac agtgggaagc tctacaaggc caagagcaac aaggagctgt atggcttcct
     2461 tttcaacgac ttcctcctgc tgactcagat cacgaagcct ttggggtctt ctggcaccga
     2521 caaagtcttc agccccaaat caaacctgca gtataaaatg tataaaacac ctattttcct
     2581 aaatgaggtt ctagtaaaat tacccaccga cccttctgga gacgagccca tcttccacat
     2641 ctcccacatt gaccgcgtct atactctccg agcagaaagc ataaatgaaa ggactgcctg
     2701 ggtgcagaaa atcaaagctg cttctgaact ctacatagag actgagaaaa agaagcgcga
     2761 gaaagcgtac ctggtccgtt cccaaagggc aacaggcatt ggaaggttga tggtgaacgt
     2821 ggttgaaggc atcgagttga aaccctgtcg gtcacatgga aagagcaacc cgtactgtga
     2881 ggtgaccatg ggttcccagt gccacatcac caagacgatc caggacactc tgaaccccaa
     2941 gtggaattcc aactgccagt tcttcatccg agacctggag caggaagtcc tctgcatcac
     3001 tgtgttcgag agggaccagt tctcaccaga tgattttttg ggtcggacgg agatccgcgt
     3061 ggcggacatc aagaaagacc agggctccaa aggtccagtt acgaagtgtc ttctgctgca
     3121 cgaagtcccc acgggagaga ttgtggtccg cttggacctg cagttgtttg atgagccgtg
     3181 tacagactac aaagaccatg acggtgatta taaagatcat gacatcgact acaaggatga
     3241 cgatgacaag taactcgaga taatcaacct ctggattaca aaatttgtga aagattgact
     3301 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
     3361 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggtta
     3421 gttcttgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg
     3481 ctgttgggca ctgacaattc cgtggtgttt atttgtgaaa tttgtgatgc tattgcttta
     3541 tttgtaacca tctagcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     3601 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     3661 gggggagatg tgggaggttt tttaaagatc tggagaccgg tgtggcacgt gcggaccgag
     3721 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     3781 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     3841 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     3901 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     3961 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     4021 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     4081 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     4141 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     4201 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     4261 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     4321 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     4381 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     4441 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4501 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4561 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     4621 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     4681 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     4741 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     4801 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     4861 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     4921 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     4981 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     5041 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     5101 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     5161 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     5221 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     5281 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     5341 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     5401 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     5461 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     5521 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     5581 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     5641 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     5701 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     5761 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     5821 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     5881 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     5941 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     6001 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     6061 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     6121 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     6181 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     6241 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     6301 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     6361 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     6421 cctttttacg gttcctggcc ttttgctggc cttttgctca catgt
//
