LOCUS Exported 6844 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Cre recombinase-dependent expression of GFP, i.e. GFP is only expressed in the presence of Cre due to a transcriptional stop in between the loxP sites.. ACCESSION . VERSION . KEYWORDS pCALNL-GFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6844) AUTHORS Matsuda T, Cepko CL TITLE Controlled expression of transgenes introduced by in vivo electroporation. JOURNAL Proc Natl Acad Sci U S A. 2007 Jan 5. ():. PUBMED 17209010 REFERENCE 2 (bases 1 to 6844) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6844 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 4..383 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 386..661 /label=chicken beta-actin promoter intron 662..1670 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 1594..1616 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 1678..1697 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" protein_bind 1745..1778 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS 1828..2622 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(1882..1901) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 2492..2511 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 2802..2936 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" protein_bind 2951..2984 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." regulatory 3056..3065 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 3062..3781 /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 3062..3778 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(3107..3128) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(3368..3387) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 3715..3736 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(3868..3887) /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" polyA_signal 3933..3988 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(3987..4006) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(4347..4363) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4347..4363) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4360..4382) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4371..4387 /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(4395..4425) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4440..4461 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 4520..4715 /label=SV40 promoter /note="SV40 early promoter" rep_origin 4566..4701 /label=SV40 ori /note="SV40 origin of replication" primer_bind 4628..4647 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" polyA_signal 4721..4855 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4923..4940) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5094..5682) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5174..5193) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5853..6713) /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 6476..6495 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6714..6818) /gene="bla" /label=AmpR promoter ORIGIN 1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata 61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc 121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag 181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac 241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg 301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg 361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc 421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca 481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg 541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt 601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc 661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc 721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc 781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag 841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg 901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg 961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc 1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg 1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc 1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc 1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg 1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct 1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg 1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc 1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt 1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg 1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg 1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca 1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attgattaat tcgagcgaac 1741 gcgtataact tcgtatagca tacattatac gaagttatct cgagtcggat ttgatctgat 1801 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct 1861 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc 1921 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc 1981 gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc 2041 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg 2101 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag 2161 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc 2221 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt 2281 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc 2341 gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc 2401 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg 2461 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag 2521 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg 2581 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg 2641 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct 2701 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 2761 gcggggatct catgctggag ttcttcgccc accccatcga taacttgttt attgcagctt 2821 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac 2881 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatcaaat 2941 ccgaacgcgt ataacttcgt atagcataca ttatacgaag ttatctcgag tcgctcggta 3001 cgatttaaat tgaattctgc agtcgacggt accgcgggcc cgggatccac cggtcgccac 3061 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga 3121 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta 3181 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac 3241 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa 3301 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt 3361 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct 3421 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca 3481 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa 3541 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc 3601 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca 3661 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt 3721 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta 3781 aagcggccgc actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa 3841 tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat 3901 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat 3961 agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt 4021 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgccatat gctggctgcc 4081 atgaacaaag gtggctataa agaggtcatc agtatatgaa acagccccct gctgtccatt 4141 ccttattcca tagaaaagcc ttgacttgag gttagatttt ttttatattt tgttttgtgt 4201 tatttttttc tttaacatcc ctaaaatttt ccttacatgt tttactagcc agatttttcc 4261 tcctctcctg actactccca gtcatagctg tccctcttct cttatgaaga tccctcgacc 4321 tgcagcccaa gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg 4381 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa 4441 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac 4501 ctgtcgtgcc agcggatccg catctcaatt agtcagcaac catagtcccg cccctaactc 4561 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa 4621 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt 4681 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct aacttgttta ttgcagctta 4741 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact 4801 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatccgctg 4861 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct 4921 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac 4981 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga 5041 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat 5101 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac 5161 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct 5221 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg 5281 ctttctcaat gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg 5341 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt 5401 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg 5461 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac 5521 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga 5581 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt 5641 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt 5701 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga 5761 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc 5821 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct 5881 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata 5941 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca 6001 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga 6061 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga 6121 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg 6181 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga 6241 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt 6301 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct 6361 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca 6421 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat 6481 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga 6541 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc 6601 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg 6661 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc 6721 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt 6781 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca 6841 cctg //