LOCUS Exported 7468 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 7468) 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..7468 /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)" promoter 2319..2417 /label=U6 consensus mosquito /label=U6 consensus mosquito 100 /note="synthetic mosquito consensus promoter " misc_RNA 2436..2511 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" misc_feature 2520..2684 /label=3'end snRNAU696AB Dmel promoter 2743..5252 /label=Ac5 promoter /note="Drosophila melanogaster actin 5C promoter" CDS 5286..6002 /codon_start=1 /product="enhanced blue variant of GFP (Ai et al., 2007)" /label=EBFP2 /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL EFRTAAGITLGMDELYK" CDS 6006..6059 /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 6084..6683 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" polyA_signal 6710..6831 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" protein_bind 7025..7058 /label=attB /bound_moiety="phage phi-C31 integrase" /note="minimal attB site for the phi-C31 integrase (Grothet al., 2000)" primer_bind complement(7074..7090) /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 acttcgaatt cccactattc aatcacttaa 2341 aaatataagt ttatccatcg ctagaaccgt tttgacgtgc gctaactata taagcaaaat 2401 ggtttccctc tatacttggg tcttcgagaa gacctgtttt agagctagaa atagcaagtt 2461 aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cttttttttt 2521 tttgcctacc tggagcctga gagttgttca ataaaataaa aatgtttcgt ttttttgctt 2581 tcgccagtat ttattatttt tcatcaatat gtattcaatt tggtatgtat ttagtaattg 2641 taatatatag acaatggttt tccgttgacg tacatacatc tgacggtacc cagcatgcaa 2701 ttctatattc taaaaacaca aatgatactt ctaaaaaaaa atcatgaatg gcatcaactc 2761 tgaatcaaat ctttgcagat gcacctactt ctcatttcca ctgtcacatc atttttccag 2821 atctcgctgc ctgttatgtg gcccacaaac caagacacgt tttatggcca ttaaagctgg 2881 ctgatcgtcg ccaaacacca aatacataat gaatatgtac acattcgaga aagaagcgat 2941 caaagaagca tcttcgggcg gagtaggaga atgcggagga gaaggagaac gagctgatct 3001 agtatctctc cacaatccaa tgccaactga ccaactggcc atattcggag caatttgaag 3061 ccaatttcca tcgcctggcg atcgctccat tcttggctat atgtttttca ccgttacccg 3121 gggccatttt caaagactcg tcggcaagat aagattgtgt cactcgctgt ctctcttcat 3181 ttgtcgaaga atgctgagga atttcgcgat gacgtcggcg agtattttga agaatgagaa 3241 taatttgtat ttatacgaaa atcagttagt ggaattttct acaaaaacat gttatctata 3301 gataattttg ttgcaaaata tgttgactat gacaaagatt gtatgtatat acctttaatg 3361 tattctcatt ttcttatgta tttataatgg caatgatgat actgatgata ttttaagatg 3421 atgccagacc aaaaggcttg aatttctgcg tcttttgccg aacgcagtgc atgtgcaatt 3481 gttgtttttt ggaatattca attttcggac tgtccgcttt gatttcagtt tcttggctta 3541 ttcaaaaagc aaagtaaagc caaaaaagcg agatggcaat accaaatgcg gcaaaacggt 3601 agtggaagga aaggggtgcg gggcagcgga aggaagggtg gggcggggcg tggcggggtc 3661 tgtggctggg cgcgacgtca ccgacgttgg agccactcct ttgaccatgt gtgcgtgtgt 3721 gtattattcg tgtctcgcca ctcgccggtt gtttttttct ttttatgctg cgctctctct 3781 agcgccatct cgcttacgca tgctcaacgc accgcatgtt gccgtttcct tttatgcgtc 3841 attttggctc gaaataggca attatttaaa caaagattag tcaacgaaaa cgctaaaata 3901 aataagtcta caatatggtt acttattgcc atgtgtgtgc agccaacgat agcaacaaaa 3961 gcaacaacac aggtggcttt ccctctttca ctttttgttt gcaagccgcg tgcgagcaag 4021 acggcacgac cggcaaacgc aattacgctg acaaagagca gacgaagttt tggcgaaaaa 4081 catcaaggcg cctgatacga atgcatttgc aataacaatt gcgatattta atattgttta 4141 tgaagctgtt tgacttcaaa acacacaaaa aaaaaaataa aacaaattat ttgaaagaga 4201 attaggaatc ggacgcttat cgttagggta acaacaagaa atgcttactg agtcacagcc 4261 tctggaaaac tgccgcaagc cagagagaga gagaaaaaga gggagagcag cttagaccgc 4321 atgtgcttgt gtgtgaggcg tctctctctt cgtctctgtt gcgcaaacgc atagactgca 4381 ctgagaaaat cgattaccta ttttttatga atgaatattt gcactattac tattcaaaac 4441 tattaagata gcaatcacat tcaatagcca aatactatac cacctgagcg atgcaacgaa 4501 atgatcaatt tgagcaaaaa tgctgcatat ttaggacggc atcattatag aaatgcttct 4561 tgctgtgtac ttttctctcg tctggcagct gtttcgccgt tattgttaaa accggcttaa 4621 gttaggtgtg ttttctacga ctagtgaatg ccctactaga agatgtgtgt tgcacaaaat 4681 gtccctggaa taaccaattt gaagtgcaga tagcagtaaa cgtaagctaa tatgaatatt 4741 atttaactgt aatgttttaa tatcgctgga cattactaat aaacccacta taaacacatg 4801 tacatatgta tgttttggca tacaatgagt agttggggaa aaaatgtgta aaagcaccgt 4861 gaccatcaca gcataaagat aaccagctga agtatcgaat atgagtaacc cccaaattga 4921 atcacatgcc gcaactgata ggacccatgg aagtacactc ttcatggcga tatacaagac 4981 acacacaagc acgaacaccc agttgcggag gaaattctcc gtaaatgaaa acccaatcgg 5041 cgaacaattc atacccatat atggtaaaag ttttgaacgc gacttgagag cggagagcat 5101 tgcggctgat aaggttttag cgctaagcgg gctttataaa acgggctgcg ggaccagttt 5161 tcatatcact accgtttgag ttcttgtgct gtgtggatac tcctcccgac acaaagccgc 5221 tccatcagcc agcagtcgtc taatccagag accccggatc cagatgaatt cgcccttgtt 5281 taaacatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc 5341 tggacggcga cgtaaacggc cacaagttca gcgtgagggg cgagggcgag ggcgatgcca 5401 ccaacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc 5461 ccaccctcgt gaccaccctg agccacggcg tgcagtgctt cgcccgctac cccgaccaca 5521 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca 5581 tcttcttcaa ggacgacggc acctacaaga cccgcgccga ggtgaagttc gagggcgaca 5641 ccctagtgaa ccgcatcgag ctgaagggcg tcgacttcaa ggaggacggc aacatcctgg 5701 ggcacaagct ggagtacaac ttcaacagcc acaacatcta tatcatggcc gtcaagcaga 5761 agaacggcat caaggtgaac ttcaagatcc gccacaacgt ggaggacggc agcgtgcagc 5821 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca 5881 gccactacct gagcacccag tccgtgctga gcaaagaccc caacgagaag cgcgatcaca 5941 tggtcctgct ggagttccgc accgccgccg ggatcactct cggcatggac gagctgtaca 6001 agcttgaggg cagaggaagc cttctaacat gcggtgacgt ggaggagaat cccggccctg 6061 ctagcggtag cggcagcggt agcatgaccg agtacaagcc cacggtgcgc ctcgccaccc 6121 gcgacgacgt cccccgggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca 6181 cgcgccacac cgtcgacccg gaccgccaca tcgagcgggt caccgagctg caagaactct 6241 tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg 6301 tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc 6361 cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc 6421 tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg 6481 accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc 6541 gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc 6601 ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca 6661 tgacccgcaa gcccggtgcc tgactcgagg gatccgaatt cagacatgat aagatacatt 6721 gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt 6781 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac 6841 aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag 6901 taaaacctct acaaatgtgg tatggctgat tatgatcagt cgaccgatgc ccttgagagc 6961 cttcaaccca gtcagctcct tccggtgggc gcggggcatg actatcgtcg ccgcacttat 7021 ggtacgcgcc cggggagccc aagggcacgc cctggcaccc gaagggcgaa ttcactggcc 7081 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca 7141 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc 7201 caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat 7261 ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca 7321 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg 7381 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg 7441 ttttcaccgt catcaccgaa acgcgcga //