LOCUS Exported 7265 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses hFMRP iso 1. ACCESSION . VERSION . KEYWORDS pFRT-TODestFLAGHAhFMRPiso1 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7265) AUTHORS Ascano M Jr, Mukherjee N, Bandaru P, Miller JB, Nusbaum JD, Corcoran DL, Langlois C, Munschauer M, Dewell S, Hafner M, Williams Z, Ohler U, Tuschl T TITLE FMRP targets distinct mRNA sequence elements to regulate protein expression. JOURNAL Nature. 2012 Dec 20;492(7429):382-6. doi: 10.1038/nature11737. Epub 2012 Dec 12. PUBMED 23235829 REFERENCE 2 (bases 1 to 7265) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7265 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..380 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 381..584 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 535..555 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" protein_bind 586..604 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 607..625 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" primer_bind 635..659 /label=LNCX /note="Human CMV promoter, forward primer" CDS 759..782 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 783..809 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" protein_bind 825..849 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" protein_bind complement(2785..2809) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" CDS 2862..2903 /codon_start=1 /product="epitope tag from simian virus 5" /label=V5 tag /translation="GKPIPNPLLGLDST" primer_bind complement(2983..3000) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 2989..3213 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" primer_bind complement(3346..3365) /label=F1ori-R /note="F1 origin, reverse primer" protein_bind 3497..3544 /label=FRT /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="FLP-mediated recombination occurs in the 8-bp core sequence TCTAGAAA (Turan and Bode, 2011)." CDS 3552..4574 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /label=HygR /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSASDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4704..4825 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4741..4760) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4795..4814 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(4874..4890) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4874..4890) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4887..4909) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4898..4914 /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(4922..4952) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4967..4988 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5105..5122) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5276..5864) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5356..5375) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6035..6895) /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 6658..6677 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6896..7000) /gene="bla" /label=AmpR promoter primer_bind complement(7075..7094) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc 61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca 121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga 181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc 241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctccct atcagtgata 601 gagatctccc tatcagtgat agagatcgtc gacgagctcg tttagtgaac cgtcagatcg 661 cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc 721 tccggactct agcgtttaaa cttaagcttg gtaccatgga ctacaaggac gacgatgaca 781 agtaccctta tgacgtgccc gattacgctt tgaaccagat atcaacaagt ttgtacaaaa 841 aagcaggctc caccatggag gagctggtgg tggaagtgcg gggctccaat ggcgctttct 901 acaaggcatt tgtaaaggat gttcatgaag attcaataac agttgcattt gaaaacaact 961 ggcagcctga taggcagatt ccatttcatg atgtcagatt cccacctcct gtaggttata 1021 ataaagatat aaatgaaagt gatgaagttg aggtgtattc cagagcaaat gaaaaagagc 1081 cttgctgttg gtggttagct aaagtgagga tgataaaggg tgagttttat gtgatagaat 1141 atgcagcatg tgatgcaact tacaatgaaa ttgtcacaat tgaacgtcta agatctgtta 1201 atcccaacaa acctgccaca aaagatactt tccataagat caagctggat gtgccagaag 1261 acttacggca aatgtgtgcc aaagaggcgg cacataagga ttttaaaaag gcagttggtg 1321 ccttttctgt aacttatgat ccagaaaatt atcagcttgt cattttgtcc atcaatgaag 1381 tcacctcaaa gcgagcacat atgctgattg acatgcactt tcggagtctg cgcactaagt 1441 tgtctctgat aatgagaaat gaagaagcta gtaagcagct ggagagttca aggcagcttg 1501 cctcgagatt tcatgaacag tttatcgtaa gagaagatct gatgggtcta gctattggta 1561 ctcatggtgc taatattcag caagctagaa aagtacctgg ggtcactgct attgatctag 1621 atgaagatac ctgcacattt catatttatg gagaggatca ggatgcagtg aaaaaagcta 1681 gaagctttct cgaatttgct gaagatgtaa tacaagttcc aaggaactta gtaggcaaag 1741 taataggaaa aaatggaaag ctgattcagg agattgtgga caagtcagga gttgtgaggg 1801 tgaggattga ggctgaaaat gagaaaaatg ttccacaaga agaggaaatt atgccaccaa 1861 attcccttcc ttccaataat tcaagggttg gacctaatgc cccagaagaa aaaaaacatt 1921 tagatataaa ggaaaacagc acccattttt ctcaacctaa cagtacaaaa gtccagaggg 1981 tgttagtggc ttcatcagtt gtagcagggg aatcccagaa acctgaactc aaggcttggc 2041 agggtatggt accatttgtt tttgtgggaa caaaggacag catcgctaat gccactgttc 2101 ttttggatta tcacctgaac tatttaaagg aagtagacca gttgcgtttg gagagattac 2161 aaattgatga gcagttgcga cagattggag ctagttctag accaccacca aatcgtacag 2221 ataaggaaaa aagctatgtg actgatgatg gtcaaggaat gggtcgaggt agtagacctt 2281 acagaaatag ggggcacggc agacgcggtc ctggatatac ttcaggaact aattctgaag 2341 catcaaatgc ttctgaaaca gaatctgacc acagagacga actcagtgat tggtcattag 2401 ctccaacaga ggaagagagg gagagcttcc tgcgcagagg agacggacgg cggcgtggag 2461 ggggaggaag aggacaagga ggaagaggac gtggaggagg cttcaaagga aacgacgatc 2521 actcccgaac agataatcgt ccacgtaatc caagagaggc taaaggaaga acaacagatg 2581 gatcccttca gatcagagtt gactgcaata atgaaaggag tgtccacact aaaacattac 2641 agaatacctc cagtgaaggt agtcggctgc gcacgggtaa agatcgtaac cagaagaaag 2701 agaagccaga cagcgtggat ggtcagcaac cactcgtgaa tggagtaccc taaacgaatt 2761 cgcggccgca ctcgagatat ctagacccag ctttcttgta caaagtggtt gatatccagc 2821 acagtggcgg ccgctcaagt ctagagggcc cgcggttcga aggtaagcct atccctaacc 2881 ctctcctcgg tctcgattct acgcgtacct gatccgcggc cgcatagata actgatccag 2941 tgtgcctcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta 3001 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca 3061 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc 3121 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata 3181 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg 3241 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg 3301 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct 3361 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc 3421 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg 3481 atggttcacg tacctagaag ttcctattcc gaagttccta ttctctagaa agtataggaa 3541 cttccttggc caaaaagcct gaactcaccg cgacgtctgt cgagaagttt ctgatcgaaa 3601 agttcgacag cgcctccgac ctgatgcagc tctcggaggg cgaagaatct cgtgctttca 3661 gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc gatggtttct 3721 acaaagatcg ttatgtttat cggcactttg catcggccgc gctcccgatt ccggaagtgc 3781 ttgacattgg ggaattcagc gagagcctga cctattgcat ctcccgccgt gcacagggtg 3841 tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg gtcgcggagg 3901 ccatggatgc gatcgctgcg gccgatctta gccagacgag cgggttcggc ccattcggac 3961 cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt gctgatcccc 4021 atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc gcgcaggctc 4081 tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc gtgcacgcgg 4141 atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc attgactgga 4201 gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc tggaggccgt 4261 ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg gagcttgcag 4321 gatcgccgcg gctccgggcg tatatgctcc gcattggtct tgaccaactc tatcagagct 4381 tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac gcaatcgtcc 4441 gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg gccgtctgga 4501 ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc actcgtccga 4561 gggcaaagga atagcacgta ctacgagatt tcgattccac cgccgccttc tatgaaaggt 4621 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca 4681 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa 4741 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt 4801 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct 4861 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac 4921 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac 4981 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc 5041 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg 5101 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc 5161 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt 5221 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc 5281 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa 5341 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc 5401 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg 5461 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc 5521 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc 5581 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca 5641 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact 5701 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg 5761 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt 5821 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct 5881 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga 5941 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa 6001 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac 6061 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga 6121 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc 6181 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca 6241 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta 6301 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg 6361 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc 6421 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg 6481 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt 6541 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt 6601 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata 6661 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc 6721 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac 6781 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa 6841 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct 6901 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat 6961 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc 7021 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat 7081 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc 7141 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat 7201 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac 7261 gcgtt //