LOCUS       62056_2555        5982 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5982)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..5982
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(462..473)
                     /codon_start=1
                     /label=Factor Xa site
                     /note="Factor Xa recognition and cleavage site"
                     /translation="IEGR"
     polyA_signal    complement(622..743)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1271..1859)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2033..2890)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(2891..2995)
                     /label=AmpR promoter
     rep_origin      complement(3021..3476)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     protein_bind    3728..3748
                     /label=attB4
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     promoter        3780..4020
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_RNA        4030..4059
                     /label=RfxCas13d DR30
                     /note="5' processed 30-base direct repeat for an RfxCas13d
                     gRNA (Konermann et al., 2018)"
     enhancer        4141..4444
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4445..4648
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     protein_bind    4695..4719
                     /label=attB1
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             4765..4785
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     CDS             4804..5832
                     /codon_start=1
                     /label=Cre
                     /note="site-specific recombinase"
                     /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     CDS             5851..5898
                     /codon_start=1
                     /label=nucleoplasmin NLS
                     /note="bipartite nuclear localization signal from
                     nucleoplasmin"
                     /translation="KRPAATKKAGQAKKKK"
     protein_bind    complement(5933..5957)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    5973..5982
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
ORIGIN
        1 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
       61 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
      121 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
      181 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
      241 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
      301 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
      361 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
      421 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
      481 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
      541 ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccgt cgacccgggc
      601 ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
      661 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
      721 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
      781 agggggagat gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcg
      841 ataaggatct tcctagagca tggctacgta gataagtagc atggcgggtt aatcattaac
      901 tacaaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact
      961 gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc
     1021 gagcgagcgc gcagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     1081 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     1141 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     1201 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     1261 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     1321 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     1381 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     1441 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     1501 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     1561 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     1621 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     1681 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     1741 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     1801 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     1861 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     1921 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     1981 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     2041 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     2101 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     2161 tgataccgcg tgacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     2221 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     2281 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     2341 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     2401 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     2461 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     2521 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     2581 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     2641 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     2701 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     2761 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     2821 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     2881 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     2941 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     3001 ttccccgaaa agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     3061 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     3121 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     3181 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     3241 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     3301 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     3361 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     3421 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca
     3481 ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt
     3541 acgccagctg cgcgctcgct cgctcactga ggccgcccgg gcaaagcccg ggcgtcgggc
     3601 gacctttggt cgcccggcct cagtgagcga gcgagcgcgc agagagggag tggccaactc
     3661 catcactagg ggttccttgt agttaatgat taacccgcca tgccggttca ccaattgtat
     3721 ccgatatcaa ctttgtatag aaaagttggc tccgaatttc tcgaggaatt cggcaggaag
     3781 agggcctatt tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga
     3841 taattagaat taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa
     3901 agtaataatt tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata
     3961 tgcttaccgt aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaaggac
     4021 gaaacaccga acccctacca actggtcggg gtttgaaact gagaccatcg gtctcatttt
     4081 tttaagctta tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     4141 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     4201 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     4261 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     4321 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     4381 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     4441 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
     4501 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
     4561 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
     4621 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc ccgggtaact
     4681 gatcataatt cgacccaagt ttgtacaaaa aagcaggctg attaccggag aattccaatt
     4741 ggcggccgca ccggtgccac catgccaaag aagaagcgga aggtcggtgg cggctcaccc
     4801 gggatgtcca atttactgac cgtacaccaa aatttgcctg cattaccggt cgatgcaacg
     4861 agtgatgagg ttcgcaagaa cctgatggac atgttcaggg atcgccaggc gttttctgag
     4921 catacctgga aaatgcttct gtccgtttgc cggtcgtggg cggcatggtg caagttgaat
     4981 aaccggaaat ggtttcccgc agaacctgaa gatgttcgcg attatcttct atatcttcag
     5041 gcgcgcggtc tggcagtaaa aactatccag caacatttgg gccagctaaa catgcttcat
     5101 cgtcggtccg ggctgccacg accaagtgac agcaatgctg tttcactggt tatgcggcgg
     5161 atccgaaaag aaaacgttga tgccggtgaa cgtgcaaaac aggctctagc gttcgaacgc
     5221 actgatttcg accaggttcg ttcactcatg gaaaatagcg atcgctgcca ggatatacgt
     5281 aatctggcat ttctggggat tgcttataac accctgttac gtatagccga aattgccagg
     5341 atcagggtta aagatatctc acgtactgac ggtgggagaa tgttaatcca tattggcaga
     5401 acgaaaacgc tggttagcac cgcaggtgta gagaaggcac ttagcctggg ggtaactaaa
     5461 ctggtcgagc gatggatttc cgtctctggt gtagctgatg atccgaataa ctacctgttt
     5521 tgccgggtca gaaaaaatgg tgttgccgcg ccatctgcca ccagccagct atcaactcgc
     5581 gccctggaag ggatttttga agcaactcat cgattgattt acggcgctaa ggatgactct
     5641 ggtcagagat acctggcctg gtctggacac agtgcccgtg tcggagccgc gcgagatatg
     5701 gcccgcgctg gagtttcaat accggagatc atgcaagctg gtggctggac caatgtaaat
     5761 attgtcatga actatatccg taacctggat agtgaaacag gggcaatggt gcgcctgctg
     5821 gaagatggcg atactagtgg tggcggctca aagcgtcctg ctgctactaa gaaagctggt
     5881 caagctaaga aaaagaaatc tagatagtga ctcgagacta gataactgat ctacccagct
     5941 ttcttgtaca aagtggctcg agctctatcg ataatcaacc tc
//
