LOCUS Exported 5169 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expression of GFP-CaaX (sequence from Hras) in mammalian cells.. ACCESSION . VERSION . KEYWORDS EGFP-CaaX(tH) in pCSDEST2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5169) TITLE New_for_revision JOURNAL Unpublished REFERENCE 2 (bases 1 to 5169) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5169 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 38..56 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 38..55 /label=SP6 /note="SP6 promoter, forward primer" protein_bind 111..135 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" regulatory 136..145 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 142..858 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 142..858 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(187..208) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(448..467) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 795..816 /label=EGFP-C /note="EGFP, forward primer" protein_bind complement(860..884) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" promoter complement(956..973) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" polyA_signal 978..1112 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" protein_bind complement(1164..1184) /gene="mutant version of attB" /label=attB3 /bound_moiety="BP Clonase(TM)" /note="core recombination site for the Gateway(R) BP reaction" promoter complement(1202..1219) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" polyA_signal 1224..1358 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1471..1491) /label=T3 /note="T3 promoter, forward primer" promoter complement(1471..1489) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(1510..1526) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1510..1526) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1523..1545) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1534..1550 /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(1558..1588) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1603..1624 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(1741..1758) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(1912..2500) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1992..2011) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(2671..3531) /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 3294..3313 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(3532..3636) /gene="bla" /label=AmpR promoter rep_origin complement(3662..4117) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3799..3820) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 4011..4030 /label=F1ori-R /note="F1 origin, reverse primer" promoter 4195..3 /label=CMV IE94 promoter /note="enhancer/promoter region of simian cytomegalovirus major immediate early transcription unit IE94" ORIGIN 1 aaccgccatt ctgcctgggg acgtcggagc aagcttgatt taggtgacac tatagaatac 61 aagctacttg ttctttttgc aggatcccat cgattcgaat tcaaggatca acaagtttgt 121 acaaaaaagc aggctgccac catggtgagc aagggcgagg agctgttcac cggggtggtg 181 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag 241 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag 301 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc 361 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac 421 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg 481 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag 541 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc 601 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag 661 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc 721 gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac 781 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc 841 atggacgagc tgtacaagca cccagctttc ttgtacaaag tgggaaagct gaaccctcct 901 gatgagagtg gccccggctg catgagctgc aagtgtgtgc tctcctgatc tagaactata 961 gtgagtcgta ttacgtagat ccagacatga taagatacat tgatgagttt ggacaaacca 1021 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat 1081 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt 1141 ttcaggttca gggggaggtg tggcaacttt attatacata gttgcactcg agcctctaga 1201 actatagtga gtcgtattac gtagatccag acatgataag atacattgat gagtttggac 1261 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg 1321 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt 1381 ttatgtttca ggttcagggg gaggtgtggg aggtttttta attcgcggcc gcggcgccaa 1441 tgcattgggc ccggtaccca gcttttgttc cctttagtga gggttaattg cgcgcttggc 1501 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa 1561 catacgagcc gggagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac 1621 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca 1681 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc 1741 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc 1801 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc 1861 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag 1921 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc 1981 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt 2041 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct 2101 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg 2161 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct 2221 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat 2281 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg 2341 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa 2401 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt 2461 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc 2521 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt 2581 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta 2641 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat 2701 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac 2761 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg 2821 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag 2881 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt 2941 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt 3001 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt 3061 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt 3121 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct 3181 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt 3241 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac 3301 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa 3361 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa 3421 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca 3481 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct 3541 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga 3601 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc 3661 taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca 3721 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag 3781 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc 3841 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc 3901 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc 3961 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa 4021 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc 4081 acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcgc 4141 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagtc gaccgccaat 4201 tcaatatggc gtatatggac tcatgccaat tcaatatggt ggatctggac ctgtgccaat 4261 tcaatatggc gtatatggac tcgtgccaat tcaatatggt ggatctggac cccagccaat 4321 tcaatatggc ggacttggca ccatgccaat tcaatatggc ggacctggca ctgtgccaac 4381 tggggagggg tctacttggc acggtgccaa gtttgaggag gggtcttggc cctgtgccaa 4441 gtccgccata ttgaattggc atggtgccaa taatggcggc catattggct atatgccagg 4501 atcaatatat aggcaatatc caatatggcc ctatgccaat atggctattg gccaggttca 4561 atactatgta ttggccctat gccatatagt attccatata tgggttttcc tattgacgta 4621 gatagcccct cccaatgggc ggtcccatat accatatatg gggcttccta ataccgccca 4681 tagccactcc cccattgacg tcaatggtct ctatatatgg tctttcctat tgacgtcata 4741 tgggcggtcc tattgacgta tatggcgcct cccccattga cgtcaattac ggtaaatggc 4801 ccgcctggct caatgcccat tgacgtcaat aggaccaccc accattgacg tcaatgggat 4861 ggctcattgc ccattcatat ccgttctcac gccccctatt gacgtcaatg acggtaaatg 4921 gcccacttgg cagtacatca atatctatta atagtaactt ggcaagtaca ttactattgg 4981 aagtacgcca gggtacattg gcagtactcc cattgacgtc aatggcggta aatggcccgc 5041 gatggctgcc aagtacatcc ccattgacgt caatggggag gggcaatgac gcaaatgggc 5101 gttccattga cgtaaatggg cggtaggcgt gcctaatggg aggtctatat aagcaatgct 5161 cgtttaggg //