LOCUS Exported 4598 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Golden Gate entry vector at position 7, LacZ, final entry vector for the assembly. ACCESSION . VERSION . KEYWORDS pGGEV_7'_XcmI-LacZ SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4598) AUTHORS Kirchmaier S, Lust K, Wittbrodt J TITLE Golden GATEway cloning--a combinatorial approach to generate fusion and recombination constructs. JOURNAL PLoS One. 2013 Oct 7;8(10):e76117. doi: 10.1371/journal.pone.0076117. PUBMED 24116091 REFERENCE 2 (bases 1 to 4598) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4598 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin complement(1..456) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(138..159) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 350..369 /label=F1ori-R /note="F1 origin, reverse primer" promoter 971..989 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" CDS complement(1027..1410) /codon_start=1 /gene="lacZ fragment" /product="LacZ-alpha fragment of beta-galactosidase" /label=lacZ-alpha /translation="MTMITPSYLGDTIEYSSYASNALGALPYGRPAGGREFTSDIEFPR PPWRPGACDVGPNSPYSESYYNSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEA RTDRPSQQLRSLNGEWTRPVAAH" primer_bind 1176..1198 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 1190..1207 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 1191..1207 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 1214..1233 /label=T7 /note="T7 promoter, forward primer" promoter 1214..1232 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" regulatory 1263..1272 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" regulatory 1267..1276 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" promoter complement(1370..1388) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(1406..1422) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1406..1422) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1419..1441) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1430..1446 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." promoter complement(1454..1484) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1499..1520 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." intron 1746..2318 /label=-beta-globin intron /note="intron from rabbit beta-globin gene" polyA_signal 2391..2525 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2415..2434) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2469..2488 /label=SV40pA-R /note="SV40 polyA, reverse primer" rep_origin complement(2848..3436) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(3607..4467) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAISMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 4230..4249 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(4468..4572) /gene="bla" /label=AmpR promoter ORIGIN 1 aaattgtaaa cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat 61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga 121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca 181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct 241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc 301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag 361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca 421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctacgca 481 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagggc gcctccccca 541 ttgacgtcaa ttacggtaaa tggcccgcct ggctcaatgc ccattgacgt caataggacc 601 acccaccatt gacgtcaatg ggatggctca ttgcccattc atatccgttc tcacgccccc 661 tattgacgtc aatgacggta aatggcccac ttggcagtac atcaatatct attaatagta 721 acttggcaag tacattacta ttggaagtac gccagggtac attggcagta ctcccattga 781 cgtcaatggc ggtaaatggc ccgcgatggc tgccaagtac atccccattg acgtcaatgg 841 ggaggggcaa tgacgcaaat gggcgttcca ttgacgtaaa tgggcggtag gcgtgcctaa 901 tgggaggtct atataagcaa tgctcgttta gggaaccgcc attctgcctg gggacgtcgg 961 agcaagcttg atttaggtga cactatagag gtctcaagtt gcggatccag cggtggcgtg 1021 gcgcgcttaa tgcgccgcta cagggcgcgt ccattcgcca ttcaggctgc gcaactgttg 1081 ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc 1141 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac 1201 ggccagtgaa ttgtaatacg actcactata gggcgaattg ggcccgacgt cgcatgctcc 1261 cggccgccat ggcggccgcg ggaattcgat atcactagtg aattcgcggc cgcctgcagg 1321 tcgaccatat gggagagctc ccaacgcgtt ggatgcatag cttgagtatt ctatagtgtc 1381 acctaaatag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 1441 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 1501 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 1561 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 1621 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc gccacgccac cgctggtacc 1681 taagtgagac cctcatcctc ccgggtaact aagtaaggat ctcgaccgat cctgagaact 1741 tcagggtgag tttggggacc cttgattgtt ctttcttttt cgctattgta aaattcatgt 1801 tatatggagg gggcaaagtt ttcagggtgt tgtttagaat gggaagatgt cccttgtatc 1861 accatggacc ctcatgataa ttttgtttct ttcactttct actctgttga caaccattgt 1921 ctcctcttat tttcttttca ttttctgtaa ctttttcgtt aaactttagc ttgcatttgt 1981 aacgaatttt taaattcact tttgtttatt tgtcagattg taagtacttt ctctaatcac 2041 ttttttttca aggcaatcag ggtatattat attgtacttc agcacagttt tagagaacaa 2101 ttgttataat taaatgataa ggtagaatat ttctgcatat aaattctggc tggcgtggaa 2161 atattcttat tggtagaaac aactacaccc tggtcatcat cctgcctttc tctttatggt 2221 tacaatgata tacactgttt gagatgagga taaaatactc tgagtccaaa ccgggcccct 2281 ctgctaacca tgttcatgcc ttcttctctt tcctacagct cctgggcaac gtgctggttg 2341 ttgtgctgtc tcatcatttt ggcaaagaat taattcctcg acggatcgta gatccagaca 2401 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct 2461 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac 2521 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg 2581 ttttttaatt cgcggccgct ctagaactag tctgcattaa tgaatcggcc aacgcgcggg 2641 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc 2701 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac 2761 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa 2821 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc ggcccccctg acgagcatca 2881 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc 2941 gttcccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata 3001 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct caatgctcac gctgtaggta 3061 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca 3121 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga 3181 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg 3241 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg 3301 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg 3361 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag 3421 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa 3481 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat 3541 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc 3601 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc 3661 atccatagtt gcctgactgc ccgtcgtgta gataactacg atacgggagg gcttaccatc 3721 tggccccagt gctgcaatga taccgcggga cccacgctca ccggctccag atttatcagc 3781 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc 3841 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt 3901 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc 3961 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgaaa 4021 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt 4081 atcactcatg cttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg 4141 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc 4201 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa 4261 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt 4321 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt 4381 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag 4441 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta 4501 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat 4561 aggggttccg cgcacatttc cccgaaaagt gccacctg //