LOCUS Exported 5429 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION express Flag-RagC (G domain only). ACCESSION . VERSION . KEYWORDS pRK5 Flag RagC (GD) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5429) AUTHORS Lawrence RE, Cho KF, Rappold R, Thrun A, Tofaute M, Kim DJ, Moldavski O, Hurley JH, Zoncu R TITLE A nutrient-induced affinity switch controls mTORC1 activation by its Rag GTPase-Ragulator lysosomal scaffold. JOURNAL Nat Cell Biol. 2018 Sep;20(9):1052-1063. doi: 10.1038/s41556-018-0148-6. Epub 2018 Jul 30. PUBMED 30061680 REFERENCE 2 (bases 1 to 5429) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5429 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 14..393 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 394..597 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 548..568 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 594..618 /label=LNCX /note="Human CMV promoter, forward primer" promoter 828..846 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 828..845 /label=SP6 /note="SP6 promoter, forward primer" primer_bind 859..878 /label=pMT2-F /note="Synthetic intron, forward primer" CDS 937..960 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" regulatory 1729..1738 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" polyA_signal 1742..1876 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1779..1798) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1833..1852 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" promoter 1945..2302 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(1968..1988) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 2153..2288 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2215..2234 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind complement(2322..2339) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(2322..2338) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2331..2353) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 2551..3006 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2638..2657) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2848..2869 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(3023..3042) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 3142..3164 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(3202..3220) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 3288..3392 /gene="bla" /label=AmpR promoter CDS 3393..4253 /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 complement(3611..3630) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4424..5012 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4913..4932 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5166..5183 /label=L4440 /note="L4440 vector, forward primer" protein_bind 5300..5321 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5336..5366 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5374..5390 /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)." primer_bind 5379..5401 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 5398..5414 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 5398..5414 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" ORIGIN 1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca 61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct 121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta 181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac 241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt 301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag 361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat 421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat 481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc 541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt 601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga 661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt 721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta 781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac 841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc 901 acctcggttc tatcgattga attcaccatg gaaatggact acaaggatga cgatgacaag 961 ggtggtggtg gtggtgtcga ctccctgcag tacggggcgg aggagacgcc cctcgccggc 1021 agttacggcg cggccgattc gtttccaaag gacttcggct acggcgtgga ggaggaggaa 1081 gaggaggcgg cggcggcggg cggaggggtt ggggcagggg caggcggtgg ctgtggtccg 1141 gggggcgctg acagctccaa gccgaggatt ctgctcatgg gactccggcg cagcggcaag 1201 tcctccatcc agaaggtggt gtttcataag atgtcaccca acgagaccct ctttttggaa 1261 agtaccaaca agatttataa ggatgacatt tccaatagct cctttgtgaa tttccagata 1321 tgggattttc ctgggcaaat ggactttttt gacccaacct ttgactatga gatgatcttc 1381 aggggaacag gagcattgat atacgtcatt gacgcacagg atgactacat ggaggcttta 1441 acaagacttc acattactgt ttctaaagcc tacaaagtta acccagacat gaattttgag 1501 gtttttattc acaaagttga tggtctgtct gatgatcaca aaatagaaac acagagggac 1561 attcatcaaa gggccaatga tgaccttgca gatgctgggc tagaaaaact ccatcttagc 1621 ttttatctga ctagtatcta tgaccattca atatttgaag cctttagtaa ggtggtgcag 1681 aaactcattc cattagcggc cgctaagtaa gtaaggatcc ccagcttggc cgccatggcc 1741 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac 1801 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc 1861 ttatcatgtc tggatcggga attaattcgg cgcagcacca tggcctgaaa taacctctga 1921 aagaggaact tggttaggta ccttctgagg cggaaagaac cagctgtgga atgtgtgtca 1981 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct 2041 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca 2101 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc 2161 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta 2221 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt 2281 tggaggccta ggcttttgca aaaagctgtt aacagcttgg cactggccgt cgttttacaa 2341 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct 2401 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc 2461 agcctgaatg gcgaatggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt 2521 tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg 2581 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2641 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2701 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2761 ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt 2821 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca 2881 accctatctc gggctattct tttgatttat aagggatttt gccgatttcg gcctattggt 2941 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta 3001 caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc 3061 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt 3121 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac 3181 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga 3241 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta 3301 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat 3361 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc 3421 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga 3481 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca 3541 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt 3601 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg 3661 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc 3721 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata 3781 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt 3841 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag 3901 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca 3961 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg 4021 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg 4081 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag 4141 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg 4201 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag 4261 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga 4321 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt 4381 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc 4441 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc 4501 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac 4561 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac 4621 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt 4681 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct 4741 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat 4801 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt 4861 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg 4921 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt 4981 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt 5041 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg 5101 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg 5161 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc 5221 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg 5281 gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac 5341 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag 5401 gaaacagcta tgacatgatt acgaattaa //