LOCUS       Exported                6003 bp ds-DNA     circular SYN 12-FEB-2020
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6003)
  AUTHORS   Trial User
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Dec 24, 2020 from SnapGene 4.3.6
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6003
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4"
     misc_feature    124..604
                     /label=4180_sp_nMSC_U1A
                     /note="color: #a6acb3"
     misc_feature    160..275
                     /label=MSC
                     /note="color: #a6acb3"
     promoter        276..532
                     /label=U1A
                     /note="color: #ffffff; direction: RIGHT"
     regulatory      533..542
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
     CDS             548..574
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /note="color: #993366"
                     /translation="PAAKRVKLD"
     CDS             575..2710
                     /codon_start=1
                     /product="N-terminal portion of Streptococcus pyogenes Cas9
                     (Zetsche et al., 2015)"
                     /label=Cas9(N)
                     /note="color: #993366"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQV"
     primer_bind     575..604
                     /label=F
                     /note="color: black; sequence: 
                     GACAAGAAGTACAGCATCGGCCTGGACATC; added: 2020-02-10"
     misc_feature    2681..3049
                     /label=4181_sp_N_intein
                     /note="color: #a6acb3"
     primer_bind     complement(2681..2710)
                     /label=R
                     /note="color: black; sequence: 
                     CACCTGGGCTTTCTGGATGTCCTCTTTAAA; added: 2020-02-10"
     CDS             2711..3016
                     /codon_start=1
                     /label=N-INTEIN
                     /note="color: #000000"
                     /translation="CLSYETEILTVEYGLLPIGKIVEKRIECTVYSVDNNGNIYTQPVA
                     QWHDRGEQEVFEYCLEDGSLIRATKDHKFMTVDGQMLPIDEIFERELDLMRVDNLPN"
     polyA_signal    3050..3257
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     repeat_region   3266..3406
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4"
     rep_origin      3481..3936
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        4218..4322
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             4323..5183
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 4323 .. 4391 / #ccffcc / signal sequence
                        2: 4392 .. 5183 / #ccffcc
                     Cleavage site after base 4391"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5354..5942
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggcctct agaggtaccg ctagcccggg tcgactagtg
      181 atcagtgtga gggagtgtaa agctggttta aacgttgttg tttggggttg aattactctt
      241 ctagaccgcg gcgcgcctcc atggatatca agcttatgga ggcggtacta tgtagatgag
      301 aattcaggag caaactggga aaagcaactg cttccaaata tttgtgattt ttacagtgta
      361 gttttggaaa aactcttagc ctaccaattc ttctaagtgt tttaaaatgt gggagccagt
      421 acacatgaag ttatagagtg ttttaatgag gcttaaatat ttaccgtaac tatgaaatgc
      481 tacgcatatc atgctgttca ggctccgtgg ccacgcaact catactaccg gtgccaccat
      541 ggctagcccc gccgccaagc gcgtgaagct ggacgacaag aagtacagca tcggcctgga
      601 catcggcacc aactctgtgg gctgggccgt gatcaccgac gagtacaagg tgcccagcaa
      661 gaaattcaag gtgctgggca acaccgaccg gcacagcatc aagaagaacc tgatcggagc
      721 cctgctgttc gacagcggcg aaacagccga ggccacccgg ctgaagagaa ccgccagaag
      781 aagatacacc agacggaaga accggatctg ctatctgcaa gagatcttca gcaacgagat
      841 ggccaaggtg gacgacagct tcttccacag actggaagag tccttcctgg tggaagagga
      901 taagaagcac gagcggcacc ccatcttcgg caacatcgtg gacgaggtgg cctaccacga
      961 gaagtacccc accatctacc acctgagaaa gaaactggtg gacagcaccg acaaggccga
     1021 cctgcggctg atctatctgg ccctggccca catgatcaag ttccggggcc acttcctgat
     1081 cgagggcgac ctgaaccccg acaacagcga cgtggacaag ctgttcatcc agctggtgca
     1141 gacctacaac cagctgttcg aggaaaaccc catcaacgcc agcggcgtgg acgccaaggc
     1201 catcctgtct gccagactga gcaagagcag acggctggaa aatctgatcg cccagctgcc
     1261 cggcgagaag aagaatggcc tgttcggaaa cctgattgcc ctgagcctgg gcctgacccc
     1321 caacttcaag agcaacttcg acctggccga ggatgccaaa ctgcagctga gcaaggacac
     1381 ctacgacgac gacctggaca acctgctggc ccagatcggc gaccagtacg ccgacctgtt
     1441 tctggccgcc aagaacctgt ccgacgccat cctgctgagc gacatcctga gagtgaacac
     1501 cgagatcacc aaggcccccc tgagcgcctc tatgatcaag agatacgacg agcaccacca
     1561 ggacctgacc ctgctgaaag ctctcgtgcg gcagcagctg cctgagaagt acaaagagat
     1621 tttcttcgac cagagcaaga acggctacgc cggctacatt gacggcggag ccagccagga
     1681 agagttctac aagttcatca agcccatcct ggaaaagatg gacggcaccg aggaactgct
     1741 cgtgaagctg aacagagagg acctgctgcg gaagcagcgg accttcgaca acggcagcat
     1801 cccccaccag atccacctgg gagagctgca cgccattctg cggcggcagg aagattttta
     1861 cccattcctg aaggacaacc gggaaaagat cgagaagatc ctgaccttcc gcatccccta
     1921 ctacgtgggc cctctggcca ggggaaacag cagattcgcc tggatgacca gaaagagcga
     1981 ggaaaccatc accccctgga acttcgagga agtggtggac aagggcgctt ccgcccagag
     2041 cttcatcgag cggatgacca acttcgataa gaacctgccc aacgagaagg tgctgcccaa
     2101 gcacagcctg ctgtacgagt acttcaccgt gtataacgag ctgaccaaag tgaaatacgt
     2161 gaccgaggga atgagaaagc ccgccttcct gagcggcgag cagaaaaagg ccatcgtgga
     2221 cctgctgttc aagaccaacc ggaaagtgac cgtgaagcag ctgaaagagg actacttcaa
     2281 gaaaatcgag tgcttcgact ccgtggaaat ctccggcgtg gaagatcggt tcaacgcctc
     2341 cctgggcaca taccacgatc tgctgaaaat tatcaaggac aaggacttcc tggacaatga
     2401 ggaaaacgag gacattctgg aagatatcgt gctgaccctg acactgtttg aggacagaga
     2461 gatgatcgag gaacggctga aaacctatgc ccacctgttc gacgacaaag tgatgaagca
     2521 gctgaagcgg cggagataca ccggctgggg caggctgagc cggaagctga tcaacggcat
     2581 ccgggacaag cagtccggca agacaatcct ggatttcctg aagtccgacg gcttcgccaa
     2641 cagaaacttc atgcagctga tccacgacga cagcctgacc tttaaagagg acatccagaa
     2701 agcccaggtg tgcctgtcct acgagacaga gatcctgaca gtggagtatg gcctgctgcc
     2761 aatcggcaag atcgtggaga agaggatcga gtgtaccgtg tactctgtgg ataacaatgg
     2821 caacatctat acacagcccg tggcacagtg gcacgatagg ggagagcagg aggtgttcga
     2881 gtattgcctg gaggacggca gcctgatcag ggcaaccaag gaccacaagt tcatgacagt
     2941 ggatggccag atgctgccca tcgacgagat tttcgagcgg gagctggacc tgatgagagt
     3001 ggataacctg cctaattgag aattcctaga gctcgctgat cagcctcgac tgtgccttct
     3061 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     3121 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     3181 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagagaat
     3241 agcaggcatg ctggggagcg gccgcaggaa cccctagtga tggagttggc cactccctct
     3301 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     3361 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg
     3421 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     3481 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     3541 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     3601 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     3661 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     3721 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     3781 gactcttgtt ccaaactgga acaacactca accctatctc gggctattct tttgatttat
     3841 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     3901 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     3961 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     4021 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     4081 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     4141 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     4201 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4261 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4321 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     4381 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     4441 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     4501 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     4561 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     4621 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     4681 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     4741 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     4801 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     4861 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     4921 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     4981 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtggaa
     5041 gccgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     5101 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     5161 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     5221 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     5281 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     5341 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     5401 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     5461 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     5521 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     5581 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     5641 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     5701 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     5761 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     5821 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     5881 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     5941 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     6001 tgt
//
