LOCUS Exported 5385 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS Cre Stoplight SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5385) AUTHORS Yang YS, Hughes TE TITLE Cre stoplight: a red/green fluorescent reporter of Cre recombinase expression in living cells. JOURNAL Biotechniques. 2001 Nov;31(5):1036, 1038, 1040-1. PUBMED 11730010 REFERENCE 2 (bases 1 to 5385) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5385 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 6..209 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 160..180 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 254..273 /label=T7 /note="T7 promoter, forward primer" promoter 254..272 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 292..325 /label=loxH /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT). The loxH variant was optimized for expression constructs but remains compatible with loxP." regulatory 355..364 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 361..1041 /codon_start=1 /product="wild-type DsRed" /label=DsRed1 /note="mammalian codon-optimized" /translation="MVRSSKNVIKEFMRFKVRMEGTVNGHEFEIEGEGEGRPYEGHNTV KLKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG VVTVTQDSSLQDGCFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIH KALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDSKLDITSHNEDYTIVEQYERTEGRH HLFL" primer_bind complement(544..564) /label=DsRed1-N /note="DsRed1, reverse primer" primer_bind 762..781 /label=mCherry-F /note="mCherry, forward primer" primer_bind 956..979 /label=DsRed1-C /note="DsRed1, forward primer" CDS 1114..1131 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(1148..1165) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 1154..1361 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(1419..1437) /label=T7 Term /note="T7 terminator, reverse primer" terminator 1433..1480 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" protein_bind 1526..1559 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." regulatory 1589..1598 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1595..2314 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 1595..2311 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1640..1661) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1901..1920) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2248..2269 /label=EGFP-C /note="EGFP, forward primer" polyA_signal 2452..2573 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2489..2508) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2543..2562 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(2622..2638) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2622..2638) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(2635..2657) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 2646..2662 /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(2670..2700) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 2715..2736 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(2853..2870) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3024..3609) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3104..3123) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(3780..4640) /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" primer_bind 4403..4422 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(4641..4745) /gene="bla" /label=AmpR promoter primer_bind complement(4820..4839) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 5011..5 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" ORIGIN 1 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg 61 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac 121 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg 181 tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc actgcttact 241 ggcttatcga aattaatacg actcactata gggagaccca agctggctag aattacctca 301 tatagcatac attatacgaa gttatcggag gaattggctc gaggaattgc ccttgccacc 361 atggtgcgct cctccaagaa cgtcatcaag gagttcatgc gcttcaaggt gcgcatggag 421 ggcaccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag 481 ggccacaaca ccgtgaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac 541 atcctgtccc cccagttcca gtacggctcc aaggtgtacg tgaagcaccc cgccgacatc 601 cccgactaca agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc 661 gaggacggcg gcgtggtgac cgtgacccaa gactcctccc tgcaggacgg ctgcttcatc 721 tacaaggtga agttcatcgg cgtgaacttc ccctccgacg gccccgtaat gcagaagaag 781 accatgggct gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc 841 gagatccaca aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagtcc 901 atctacatgg ccaagaagcc cgtgcagctg cccggctact actacgtgga ctccaagctg 961 gacatcacct cccacaacga ggactacacc atcgtggagc agtacgagcg caccgagggc 1021 cgccaccacc tgttcctgta gcggcctcga ctaagggcaa ttcgggagct cggtaagcct 1081 atccctaacc ctctcctcgg tctcgattct agccatcatc accatcacca ttgaagctcg 1141 ctatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg 1201 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt 1261 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc 1321 aagggggagg attgggaaga caatagcagg catgctgggg attctagaag atccggctgc 1381 taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata 1441 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc 1501 cggatatccc agcggccgcg gtaccataac ttcgtatagc atacattata cgaagttatg 1561 tttcggaatt cggcttcgag atctggtcgc caccatggtg agcaagggcg aggagctgtt 1621 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag 1681 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg 1741 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt 1801 gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat 1861 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac 1921 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat 1981 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca 2041 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg 2101 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat 2161 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag 2221 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg 2281 gatcactctc ggcatggacg agctgtacaa gtaaagcggc cagatctcta gaagccgaat 2341 tcgattccac ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc 2401 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt 2461 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca 2521 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc 2581 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg 2641 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa 2701 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct 2761 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga 2821 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc 2881 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa 2941 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt 3001 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa 3061 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt 3121 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg 3181 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc 3241 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc 3301 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta 3361 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct 3421 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc 3481 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa 3541 caaaccaccg ctggtagcgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa 3601 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac 3661 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta 3721 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt 3781 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata 3841 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc 3901 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac 3961 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag 4021 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac 4081 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc 4141 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg 4201 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc 4261 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct 4321 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc 4381 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc 4441 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc 4501 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc 4561 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca 4621 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt 4681 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt 4741 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc 4801 ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc 4861 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa 4921 ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc 4981 ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt 5041 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta 5101 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga 5161 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt 5221 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta 5281 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg 5341 actttcctac ttggcagtac atctacgtat tagtcatcgc tatta //