LOCUS Exported 7091 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pT3-myr-AKT-HA SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7091) AUTHORS Calvisi DF, Wang C, Ho C, Ladu S, Lee SA, Mattu S, Destefanis G, Delogu S, Zimmermann A, Ericsson J, Brozzetti S, Staniscia T, Chen X, Dombrowski F, Evert M TITLE Increased lipogenesis, induced by AKT-mTORC1-RPS6 signaling, promotes development of human hepatocellular carcinoma. JOURNAL Gastroenterology. 2011 Mar;140(3):1071-83. Epub 2010 Dec 11. PUBMED 21147110 REFERENCE 2 (bases 1 to 7091) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7091 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 1..1179 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 232..1170 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 1127..1147 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" primer_bind 1196..1215 /label=T7 /note="T7 promoter, forward primer" promoter 1196..1214 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 1293..1317 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" CDS 1338..1358 /codon_start=1 /product="N-myristoylation signal from Src kinase (Pellman et al., 1985; Kaplan et al., 1988)" /label=myr /translation="MGSSKSK" CDS 2811..2837 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" protein_bind complement(2857..2881) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" CDS 2934..2975 /codon_start=1 /product="epitope tag from simian virus 5" /label=V5 tag /translation="GKPIPNPLLGLDST" primer_bind complement(3013..3030) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3019..3243 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" protein_bind complement(3313..3346) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind 3402..3422 /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" primer_bind complement(3804..3821) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(3804..3820) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3813..3835) /label=M13/pUC Forward /note="In lacZ gene" primer_bind complement(4029..4048) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4148..4170 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4208..4226) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4294..4398 /gene="bla" /label=AmpR promoter CDS 4399..5259 /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(4617..4636) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5430..6018 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5919..5938 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6172..6189 /label=L4440 /note="L4440 vector, forward primer" protein_bind 6306..6327 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 6342..6372 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6380..6396 /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 6385..6407 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 6404..6420 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 6404..6420 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(6846..6865) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" protein_bind complement(6918..6951) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." ORIGIN 1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg 61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt 121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca 181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc 241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt 301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg 361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg 421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct 481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg 541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc 601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga 661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct 721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca 781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg 841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg 901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat 961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg 1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa 1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca 1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata 1201 cgactcacta tagggagacc caagctggct aggtaagctt ggtaccgagc tcggatccac 1261 tagtccagtg tggtggaatt ctgcagatat caacaagttt gtacaaaaaa gcaggctccg 1321 cggccgcccc cttcaccatg gggagcagca agagcaagcc caagtctaga atgaacgacg 1381 tagccattgt gaaggagggc tggctgcaca aacgagggga atatattaaa acctggcggc 1441 cacgctactt cctcctcaag aacgatggca cctttattgg ctacaaggaa cggcctcagg 1501 atgtggatca gcgagagtcc ccactcaaca acttctcagt ggcacaatgc cagctgatga 1561 agacagagcg gccaaggccc aacaccttta tcatccgctg cctgcagtgg accacagtca 1621 ttgagcgcac cttccatgtg gaaacgcctg aggagcggga agaatgggcc accgccattc 1681 agactgtggc cgatggactc aagaggcagg aagaagagac gatggacttc cgatcaggct 1741 cacccagtga caactcaggg gctgaagaga tggaggtgtc cctggccaag cccaagcacc 1801 gtgtgaccat gaacgagttt gagtacctga aactactggg caagggcacc tttgggaaag 1861 tgattctggt gaaagagaag gccacaggcc gctactatgc catgaagatc ctcaagaagg 1921 aggtcatcgt cgccaaggat gaggttgccc acacgcttac tgagaaccgt gtcctgcaga 1981 actctaggca tcccttcctt acggccctca agtactcatt ccagacccac gaccgcctct 2041 gctttgtcat ggagtatgcc aacgggggcg agctcttctt ccacctgtct cgagagcgcg 2101 tgttctccga ggaccgggcc cgcttctatg gtgcggagat tgtgtctgcc ctggactact 2161 tgcactccga gaagaacgtg gtgtaccggg acctgaagct ggagaacctc atgctggaca 2221 aggacgggca catcaagata acggacttcg ggctgtgcaa ggaggggatc aaggatggtg 2281 ccactatgaa gacattctgc ggaacgccgg agtacctggc ccctgaggtg ctggaggaca 2341 acgactacgg ccgtgcagtg gactggtggg ggctgggcgt ggtcatgtat gagatgatgt 2401 gtggccgcct gcccttctac aaccaggacc acgagaagct gttcgagctg atcctcatgg 2461 aggagatccg cttcccgcgc acactcggcc ctgaggccaa gtccctgctc tccgggctgc 2521 tcaagaagga ccctacacag aggctcggtg ggggctctga ggatgccaag gagatcatgc 2581 agcaccggtt ctttgccaac atcgtgtggc aggatgtgta tgagaagaag ctgagcccac 2641 ctttcaagcc ccaggtcacc tctgagactg acaccaggta tttcgatgag gagttcacag 2701 ctcagatgat caccatcacg ccgcctgatc aagatgacag catggagtgt gtggacagtg 2761 agcggaggcc gcacttcccc cagttctcct actcagccag tggcacagcc tacccatacg 2821 acgtcccaga ctacgcttga aagggtgggc gcgccgaccc agctttcttg tacaaagtgg 2881 ttgatatcca gcacagtggc ggccgctcga gtctagaggg cccgcggttc gaaggtaagc 2941 ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac cggttagtaa tgagtttaaa 3001 cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc 3061 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg 3121 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg 3181 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta 3241 tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta 3301 gcggcgcatt aaataacttc gtataatgta tgctatacga agttatatcg tatgcatgct 3361 ttgcatactt ctgcctgctg gggagcctgg ggactttcca caccctaact gacacacatt 3421 ccacagaatt ccttgaaata catccacagg tacacctcca attgactcaa atgatgtcaa 3481 ttagtctatc agaagcttct aaagccatga catcattttc tggaattttc caagctgttt 3541 aaaggcacag tcaacttagt gtatgtaaac ttctgaccca ctggaattgt gatacagtga 3601 attataagtg aaataatctg tctgtaaaca attgttggaa aaatgacttg tgtcatgcac 3661 aaagtagatg tcctaactga cttgccaaaa ctattgtttg ttaacaagaa atttgtggag 3721 tagttgaaaa acgagtttta atgactccaa cttaagtgta tgtaaacttc cgacttcaac 3781 tgtagggtac cgagctcgaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac 3841 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat 3901 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg 3961 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatggtgc 4021 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca 4081 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg 4141 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga 4201 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct 4261 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc 4321 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa 4381 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt 4441 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct 4501 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc 4561 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta 4621 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac 4681 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc 4741 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac 4801 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg 4861 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac 4921 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc 4981 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt 5041 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga 5101 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc 5161 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag 5221 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca 5281 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc 5341 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca 5401 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc 5461 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta 5521 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt 5581 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc 5641 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg 5701 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg 5761 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag 5821 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc 5881 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat 5941 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg 6001 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc 6061 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt 6121 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca 6181 gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg 6241 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac 6301 gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg 6361 gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac 6421 catgattacg ccaagcttgc atgcctgcag gtcgactcta gacctataca gttgaagtcg 6481 gaagtttaca tacacttaag ttggagtcat taaaactcgt ttttcaacta ctccacaaat 6541 ttcttgttaa caaacaatag ttttggcaag tcagttagga catctacttt gtgcatgaca 6601 caagtcattt ttccaacaat tgtttacaga cagattattt cacttataat tcactgtatc 6661 acaattccag tgggtcagaa gtttacatac actaagttga ctgtgccttt aaacagcttg 6721 gaaaattcca gaaaatgatg tcatggcttt agaagcttct gatagactaa ttgacatcat 6781 ttgagtcaat tggaggtgta cctgtggatg tatttcaagg ggatccagac atgataagat 6841 acattgatga gtttggacaa accacaacta gaatgcagcg gccgctagct cgagatctta 6901 attaaactag tatcgatata acttcgtata atgtatgcta tacgaagtta tatgtacggg 6961 ccagatatac gcgttgacat tgattattga ctaggctttt gcaaaaagct ttgcaaagat 7021 ggataaagtt ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag 7081 gagtgggaat t //