LOCUS Exported 6685 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pRK5-HA GST RagC wt SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6685) AUTHORS Sancak Y, Peterson TR, Shaul YD, Lindquist RA, Thoreen CC, Bar-Peled L, Sabatini DM TITLE The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1. JOURNAL Science. 2008 Jun 13. 320(5882):1496-501. PUBMED 18497260 REFERENCE 2 (bases 1 to 6685) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6685 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 20..608 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 509..528 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 762..779 /label=L4440 /note="L4440 vector, forward primer" protein_bind 896..917 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 932..962 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 970..986 /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 975..997 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 994..1010 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 994..1010 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" enhancer 1039..1418 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1419..1622 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 1573..1593 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 1619..1643 /label=LNCX /note="Human CMV promoter, forward primer" promoter 1853..1871 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 1853..1870 /label=SP6 /note="SP6 promoter, forward primer" primer_bind 1884..1903 /label=pMT2-F /note="Synthetic intron, forward primer" regulatory 1988..1997 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2000..2026 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 2057..2710 /codon_start=1 /product="glutathione S-transferase from Schistosoma japonicum" /label=GST /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK" primer_bind 2668..2690 /label=pGEX 5' /note="pGEX vectors, Glutathione-S-transferase, forward primer" regulatory 4010..4019 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" polyA_signal 4023..4157 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4060..4079) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4114..4133 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" promoter 4226..4583 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(4249..4269) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 4434..4569 /label=SV40 ori /note="SV40 origin of replication" primer_bind 4496..4515 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind complement(4603..4620) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(4603..4619) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4612..4634) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 4832..5287 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4919..4938) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 5129..5150 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(5304..5323) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 5423..5445 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(5483..5501) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5569..5673 /gene="bla" /label=AmpR promoter CDS 5674..6534 /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(5892..5911) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" ORIGIN 1 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa 61 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt 121 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc 181 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa 241 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa 301 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc 361 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa 421 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa 481 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg 541 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc 601 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg 661 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg 721 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg 781 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat 841 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg 901 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt 961 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac atgattacga 1021 attaattcga gctcgcccga cattgattat tgactagtta ttaatagtaa tcaattacgg 1081 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc 1141 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca 1201 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg 1261 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg 1321 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt 1381 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca 1441 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg 1501 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact 1561 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag 1621 ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata 1681 gaagacaccg ggaccgatcc agcctccgcg gccgggaacg gtgcattgga acgcggattc 1741 cccgtgccaa gagtgacgta agtaccgcct atagagtcta taggcccacc cccttggctt 1801 cgttagaacg cggctacaat taatacataa ccttatgtat catacacata cgatttaggt 1861 gacactatag aataacatcc actttgcctt tctctccaca ggtgtccact cccaggtcca 1921 actgcacctc ggttctatcg attgaattcc cgaaccgaca gtcggtctct tcaccaaggc 1981 cattcgcgcc accatgggct acccctatga tgtgcctgac tacgcagatc tcaatggtgg 2041 tggtggtggg tcgagaatgt cccctatact aggttattgg aaaattaagg gccttgtgca 2101 acccactcga cttcttttgg aatatcttga agaaaaatat gaagagcatt tgtatgagcg 2161 cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct 2221 tccttattat attgatggtg atgttaaatt aacacagtct atggccatca tacgttatat 2281 agctgacaag cacaacatgt tgggtggttg tccaaaagag cgtgcagaga tttcaatgct 2341 tgaaggagcg gttttggata ttagatacgg tgtttcgaga attgcatata gtaaagactt 2401 tgaaactctc aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga 2461 tcgtttatgt cataaaacat atttaaatgg tgatcatgta acccatcctg acttcatgtt 2521 gtatgacgct cttgatgttg ttttatacat ggacccaatg tgcctggatg cgttcccaaa 2581 attagtttgt tttaaaaaac gtattgaagc tatcccacaa attgataagt acttgaaatc 2641 cagcaagtat atagcatggc ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca 2701 tcctccaaaa ggtggaggtg gtgggtcgag gggaggttct ggtggatcag gaggttcagg 2761 tggatctggg tcgacgatgt ccctgcagta cggggcggag gagacgcccc tcgccggcag 2821 ttacggcgcg gccgattcgt ttccaaagga cttcggctac ggcgtggagg aggaggaaga 2881 ggaggcggcg gcggcgggcg gaggggttgg ggcaggggca ggcggtggct gtggtccggg 2941 gggcgctgac agctccaagc cgaggattct gctcatggga ctccggcgca gcggcaagtc 3001 ctccatccag aaggtggtgt ttcataagat gtcacccaac gagaccctct ttttggaaag 3061 taccaacaag atttataagg atgacatttc caatagctcc tttgtgaatt tccagatatg 3121 ggattttcct gggcaaatgg acttttttga cccaaccttt gactatgaga tgatcttcag 3181 gggaacagga gcattgatat acgtcattga cgcacaggat gactacatgg aggctttaac 3241 aagacttcac attactgttt ctaaagccta caaagttaac ccagacatga attttgaggt 3301 ttttattcac aaagttgatg gtctgtctga tgatcacaaa atagaaacac agagggacat 3361 tcatcaaagg gccaatgatg accttgcaga tgctgggcta gaaaaactcc atcttagctt 3421 ttatctgact agtatctatg accattcaat atttgaagcc tttagtaagg tggtgcagaa 3481 actcattcca caactgccga ccttggaaaa cctattaaat atctttatat caaattcagg 3541 tattgaaaaa gcttttctct ttgatgttgt cagcaaaatc tacattgcaa cagacagttc 3601 ccctgtggat atgcaatctt atgaactttg ctgtgacatg atcgatgttg taattgatgt 3661 gtcttgtata tatgggttaa aggaagatgg aagtggaagt gcttatgaca aagaatctat 3721 ggcaattatc aagctgaata atacaactgt cctttattta aaggaggtga ctaaattttt 3781 ggcactggtc tgcattctaa gggaagaaag ctttgaaaga aaaggtttaa tagactacaa 3841 cttccactgt ttccgaaaag ctattcatga ggtttttgag gtgggtgtga cttctcacag 3901 gagctgtggt caccagacta gtgcctccag tctgaaagcg ctgacacaca atggcacgcc 3961 acgaaacgcc atctaggcgg ccgctaagta agtaaggatc cccagcttgg ccgccatggc 4021 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca 4081 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat 4141 cttatcatgt ctggatcggg aattaattcg gcgcagcacc atggcctgaa ataacctctg 4201 aaagaggaac ttggttaggt accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc 4261 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc 4321 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc 4381 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc 4441 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt 4501 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt 4561 ttggaggcct aggcttttgc aaaaagctgt taacagcttg gcactggccg tcgttttaca 4621 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc 4681 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg 4741 cagcctgaat ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat 4801 ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg 4861 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct 4921 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta 4981 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa 5041 cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct 5101 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc 5161 aaccctatct cgggctattc ttttgattta taagggattt tgccgatttc ggcctattgg 5221 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt 5281 acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc 5341 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct 5401 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca 5461 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg 5521 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct 5581 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga 5641 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc 5701 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg 5761 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc 5821 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact 5881 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc 5941 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag 6001 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat 6061 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt 6121 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa 6181 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc 6241 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg 6301 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt 6361 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca 6421 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat 6481 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca 6541 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg 6601 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg 6661 ttccactgag cgtcagaccc cgtag //