LOCUS       Exported                7685 bp ds-DNA     circular SYN 10-SEP-2021
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7685)
  AUTHORS   Enzo Mameli
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Sep 10, 2021 from SnapGene 5.1.7
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7685
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             201..1061
                     /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      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    2108..2129
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the 
                     presenceof cAMP."
     promoter        2144..2174
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator 
                     toinhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2276..2309
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Grothet
                     al., 2000)"
     misc_RNA        2594..2669
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2960..5469
                     /label=Ac5 promoter
                     /note="Drosophila melanogaster actin 5C promoter"
     CDS             5503..6219
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKGD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             6223..6276
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus 
                     capsidprotein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             6301..6900
                     /codon_start=1
                     /gene="pac from Streptomyces"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVECPKDRATWCMTRKPGA"
     polyA_signal    6927..7048
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    7242..7275
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Grothet
                     al., 2000)"
     primer_bind     complement(7291..7307)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagctt gcatgcctgc aggtcgactc tagaggatcc cccgggtgcc
     2281 agggcgtgcc cttgggctcc ccgggcgcgt acttcgaaaa tttttgaata gcccaggttg
     2341 ataaaaattc atttcatacg ttttataact tatgccccta agtatttttt gaccatagtg
     2401 tttcaattct acattaattt tacagagtag aatgaaacgc cacctactca gccaagaggc
     2461 gaaaaggtta gctcgccaag cagagagggc gccagtgctc actacttttt ataattctca
     2521 acttcttttt ccagactcag ttcgtatata tagacctatt ttcaatttaa cgtcggggtc
     2581 ttcgagaaga cctgttttag agctagaaat agcaagttaa aataaggcta gtccgttatc
     2641 aacttgaaaa agtggcaccg agtcggtgct tttttgccta cctggagcct gagagttgtt
     2701 caataaaata aaaatgtttc gtttttttgc tttcgccagt atttattatt tttcatcaat
     2761 atgtattcaa tttggtatgt atttagtaat tgtaatatat agacaatggt tttccgttga
     2821 cgtacataca tctgacgtgt gtttatttag acataatagt tatgttttca catcttttta
     2881 atgttcgctt aatgcgtatg cggtacccag catgcaattc tatattctaa aaacacaaat
     2941 gatacttcta aaaaaaaatc atgaatggca tcaactctga atcaaatctt tgcagatgca
     3001 cctacttctc atttccactg tcacatcatt tttccagatc tcgctgcctg ttatgtggcc
     3061 cacaaaccaa gacacgtttt atggccatta aagctggctg atcgtcgcca aacaccaaat
     3121 acataatgaa tatgtacaca ttcgagaaag aagcgatcaa agaagcatct tcgggcggag
     3181 taggagaatg cggaggagaa ggagaacgag ctgatctagt atctctccac aatccaatgc
     3241 caactgacca actggccata ttcggagcaa tttgaagcca atttccatcg cctggcgatc
     3301 gctccattct tggctatatg tttttcaccg ttacccgggg ccattttcaa agactcgtcg
     3361 gcaagataag attgtgtcac tcgctgtctc tcttcatttg tcgaagaatg ctgaggaatt
     3421 tcgcgatgac gtcggcgagt attttgaaga atgagaataa tttgtattta tacgaaaatc
     3481 agttagtgga attttctaca aaaacatgtt atctatagat aattttgttg caaaatatgt
     3541 tgactatgac aaagattgta tgtatatacc tttaatgtat tctcattttc ttatgtattt
     3601 ataatggcaa tgatgatact gatgatattt taagatgatg ccagaccaaa aggcttgaat
     3661 ttctgcgtct tttgccgaac gcagtgcatg tgcaattgtt gttttttgga atattcaatt
     3721 ttcggactgt ccgctttgat ttcagtttct tggcttattc aaaaagcaaa gtaaagccaa
     3781 aaaagcgaga tggcaatacc aaatgcggca aaacggtagt ggaaggaaag gggtgcgggg
     3841 cagcggaagg aagggtgggg cggggcgtgg cggggtctgt ggctgggcgc gacgtcaccg
     3901 acgttggagc cactcctttg accatgtgtg cgtgtgtgta ttattcgtgt ctcgccactc
     3961 gccggttgtt tttttctttt tatgctgcgc tctctctagc gccatctcgc ttacgcatgc
     4021 tcaacgcacc gcatgttgcc gtttcctttt atgcgtcatt ttggctcgaa ataggcaatt
     4081 atttaaacaa agattagtca acgaaaacgc taaaataaat aagtctacaa tatggttact
     4141 tattgccatg tgtgtgcagc caacgatagc aacaaaagca acaacacagg tggctttccc
     4201 tctttcactt tttgtttgca agccgcgtgc gagcaagacg gcacgaccgg caaacgcaat
     4261 tacgctgaca aagagcagac gaagttttgg cgaaaaacat caaggcgcct gatacgaatg
     4321 catttgcaat aacaattgcg atatttaata ttgtttatga agctgtttga cttcaaaaca
     4381 cacaaaaaaa aaaataaaac aaattatttg aaagagaatt aggaatcgga cgcttatcgt
     4441 tagggtaaca acaagaaatg cttactgagt cacagcctct ggaaaactgc cgcaagccag
     4501 agagagagag aaaaagaggg agagcagctt agaccgcatg tgcttgtgtg tgaggcgtct
     4561 ctctcttcgt ctctgttgcg caaacgcata gactgcactg agaaaatcga ttacctattt
     4621 tttatgaatg aatatttgca ctattactat tcaaaactat taagatagca atcacattca
     4681 atagccaaat actataccac ctgagcgatg caacgaaatg atcaatttga gcaaaaatgc
     4741 tgcatattta ggacggcatc attatagaaa tgcttcttgc tgtgtacttt tctctcgtct
     4801 ggcagctgtt tcgccgttat tgttaaaacc ggcttaagtt aggtgtgttt tctacgacta
     4861 gtgaatgccc tactagaaga tgtgtgttgc acaaaatgtc cctggaataa ccaatttgaa
     4921 gtgcagatag cagtaaacgt aagctaatat gaatattatt taactgtaat gttttaatat
     4981 cgctggacat tactaataaa cccactataa acacatgtac atatgtatgt tttggcatac
     5041 aatgagtagt tggggaaaaa atgtgtaaaa gcaccgtgac catcacagca taaagataac
     5101 cagctgaagt atcgaatatg agtaaccccc aaattgaatc acatgccgca actgatagga
     5161 cccatggaag tacactcttc atggcgatat acaagacaca cacaagcacg aacacccagt
     5221 tgcggaggaa attctccgta aatgaaaacc caatcggcga acaattcata cccatatatg
     5281 gtaaaagttt tgaacgcgac ttgagagcgg agagcattgc ggctgataag gttttagcgc
     5341 taagcgggct ttataaaacg ggctgcggga ccagttttca tatcactacc gtttgagttc
     5401 ttgtgctgtg tggatactcc tcccgacaca aagccgctcc atcagccagc agtcgtctaa
     5461 tccagagacc ccggatccag atgaattcgc ccttgtttaa acatggtgag caagggcgag
     5521 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     5581 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     5641 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
     5701 tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag
     5761 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaaggg cgacggcaac
     5821 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     5881 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     5941 aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc
     6001 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
     6061 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     6121 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     6181 gccgccggga tcactctcgg catggacgag ctgtacaagc ttgagggcag aggaagcctt
     6241 ctaacatgcg gtgacgtgga ggagaatccc ggccctgcta gcggtagcgg cagcggtagc
     6301 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc ccgggccgta
     6361 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgacccggac
     6421 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     6481 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     6541 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     6601 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     6661 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     6721 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     6781 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     6841 gacgtcgagt gcccgaagga ccgcgcgacc tggtgcatga cccgcaagcc cggtgcctga
     6901 ctcgagggat ccgaattcag acatgataag atacattgat gagtttggac aaaccacaac
     6961 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     7021 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     7081 ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat
     7141 ggctgattat gatcagtcga ccgatgccct tgagagcctt caacccagtc agctccttcc
     7201 ggtgggcgcg gggcatgact atcgtcgccg cacttatggt acgcgcccgg ggagcccaag
     7261 ggcacgccct ggcacccgaa gggcgaattc actggccgtc gttttacaac gtcgtgactg
     7321 ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg
     7381 gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg
     7441 cgaatggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     7501 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     7561 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     7621 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     7681 cgcga
//
