LOCUS       sequence-70560-e        7307 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses hFMRP iso 1 I304N.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7307)
  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 7307)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/nature11737"; journalName: "Nature"; date: "2012-12-20- 
            20"; volume: "492"; issue: "7429"; pages: "382-6"
COMMENT     Sequence Label: pFRT-TODestFLAGHAhFMRPiso1I304N
FEATURES             Location/Qualifiers
     source          1..7307
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..42
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(86..105)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        277..656
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        657..860
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     811..831
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    862..880
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    883..901
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     911..935
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1035..1058
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1059..1085
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1101..1125
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3061..3085)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3138..3179
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3259..3276)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3265..3489
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3622..3641)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3773..3820
                     /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             3828..4850
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3828..4850
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4980..5101
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5017..5036)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5071..5090
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5150..5166)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5150..5166)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5163..5185)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5174..5190
                     /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(join(5198..5204,5205..5222,5223..5228))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5243..5264
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5381..5398)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5552..6140)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5632..5651)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6311..7102,7103..7171))
                     /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     6934..6953
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7172..7276)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtaagccta tccctaaccc tctcctcggt ctcgattcta cggacggatc gggagatctc
       61 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
      121 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
      181 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
      241 cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt
      301 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      361 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      421 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      481 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      541 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
      601 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
      661 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
      721 gtctccaccc cattgacgtc aatgggagtt tgttttggaa ccaaaatcaa cgggactttc
      781 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
      841 aggtctatat aagcagagct ctccctatca gtgatagaga tctccctatc agtgatagag
      901 atcgtcgacg agctcgttta gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt
      961 ttgacctcca tagaagacac cgggaccgat ccagcctccg gactctagcg tttaaactta
     1021 agcttggtac catggactac aaggacgacg atgacaagta cccttatgac gtgcccgatt
     1081 acgctttgaa ccagatatca acaagtttgt acaaaaaagc aggctccacc atggaggagc
     1141 tggtggtgga agtgcggggc tccaatggcg ctttctacaa ggcatttgta aaggatgttc
     1201 atgaagattc aataacagtt gcatttgaaa acaactggca gcctgatagg cagattccat
     1261 ttcatgatgt cagattccca cctcctgtag gttataataa agatataaat gaaagtgatg
     1321 aagttgaggt gtattccaga gcaaatgaaa aagagccttg ctgttggtgg ttagctaaag
     1381 tgaggatgat aaagggtgag ttttatgtga tagaatatgc agcatgtgat gcaacttaca
     1441 atgaaattgt cacaattgaa cgtctaagat ctgttaatcc caacaaacct gccacaaaag
     1501 atactttcca taagatcaag ctggatgtgc cagaagactt acggcaaatg tgtgccaaag
     1561 aggcggcaca taaggatttt aaaaaggcag ttggtgcctt ttctgtaact tatgatccag
     1621 aaaattatca gcttgtcatt ttgtccatca atgaagtcac ctcaaagcga gcacatatgc
     1681 tgattgacat gcactttcgg agtctgcgca ctaagttgtc tctgataatg agaaatgaag
     1741 aagctagtaa gcagctggag agttcaaggc agcttgcctc gagatttcat gaacagttta
     1801 tcgtaagaga agatctgatg ggtctagcta ttggtactca tggtgctaat attcagcaag
     1861 ctagaaaagt acctggggtc actgctattg atctagatga agatacctgc acatttcata
     1921 tttatggaga ggatcaggat gcagtgaaaa aagctagaag ctttctcgaa tttgctgaag
     1981 atgtaataca agttccaagg aacttagtag gcaaagtaat aggaaaaaat ggaaagctga
     2041 atcaggagat tgtggacaag tcaggagttg tgagggtgag gattgaggct gaaaatgaga
     2101 aaaatgttcc acaagaagag gaaattatgc caccaaattc ccttccttcc aataattcaa
     2161 gggttggacc taatgcccca gaagaaaaaa aacatttaga tataaaggaa aacagcaccc
     2221 atttttctca acctaacagt acaaaagtcc agagggtgtt agtggcttca tcagttgtag
     2281 caggggaatc ccagaaacct gaactcaagg cttggcaggg tatggtacca tttgtttttg
     2341 tgggaacaaa ggacagcatc gctaatgcca ctgttctttt ggattatcac ctgaactatt
     2401 taaaggaagt agaccagttg cgtttggaga gattacaaat tgatgagcag ttgcgacaga
     2461 ttggagctag ttctagacca ccaccaaatc gtacagataa ggaaaaaagc tatgtgactg
     2521 atgatggtca aggaatgggt cgaggtagta gaccttacag aaataggggg cacggcagac
     2581 gcggtcctgg atatacttca ggaactaatt ctgaagcatc aaatgcttct gaaacagaat
     2641 ctgaccacag agacgaactc agtgattggt cattagctcc aacagaggaa gagagggaga
     2701 gcttcctgcg cagaggagac ggacggcggc gtggaggggg aggaagagga caaggaggaa
     2761 gaggacgtgg aggaggcttc aaaggaaacg acgatcactc ccgaacagat aatcgtccac
     2821 gtaatccaag agaggctaaa ggaagaacaa cagatggatc ccttcagatc agagttgact
     2881 gcaataatga aaggagtgtc cacactaaaa cattacagaa tacctccagt gaaggtagtc
     2941 ggctgcgcac gggtaaagat cgtaaccaga agaaagagaa gccagacagc gtggatggtc
     3001 agcaaccact cgtgaatgga gtaccctaaa cgaattcgcg gccgcactcg agatatctag
     3061 acccagcttt cttgtacaaa gtggttgata tccagcacag tggcggccgc tcaagtctag
     3121 agggcccgcg gttcgaaggt aagcctatcc ctaaccctct cctcggtctc gattctacgc
     3181 gtaccggatc cgcggccgca tagataactg atccagtgtg cctcgagtct agagggcccg
     3241 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     3301 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     3361 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     3421 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     3481 gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc
     3541 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     3601 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     3661 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     3721 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtacc tagaagttcc
     3781 tattccgaag ttcctattct ctagaaagta taggaacttc cttggccaaa aagcctgaac
     3841 tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtc tccgacctga
     3901 tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga gggcgtggat
     3961 atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa agatcgttat gtttatcggc
     4021 actttgcatc ggccgcgctc ccgattccgg aagtgcttga cattggggaa ttcagcgaga
     4081 gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa
     4141 ccgaactgcc cgctgttctg cagccggtcg cggaggccat ggatgcgatc gctgcggccg
     4201 atcttagcca gacgagcggg ttcggcccat tcggaccgca aggaatcggt caatacacta
     4261 catggcgtga tttcatatgc gcgattgctg atccccatgt gtatcactgg caaactgtga
     4321 tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg ctttgggccg
     4381 aggactgccc cgaagtccgg cacctcgtgc acgcggattt cggctccaac aatgtcctga
     4441 cggacaatgg ccgcataaca gcggtcattg actggagcga ggcgatgttc ggggattccc
     4501 aatacgaggt cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga
     4561 cgcgctactt cgagcggagg catccggagc ttgcaggatc gccgcggctc cgggcgtata
     4621 tgctccgcat tggtcttgac caactctatc agagcttggt tgacggcaat ttcgatgatg
     4681 cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc
     4741 gtacacaaat cgcccgcaga agcgcggccg tctggaccga tggctgtgta gaagtactcg
     4801 ccgatagtgg aaaccgacgc cccagcactc gtccgagggc aaaggaatag cacgtactac
     4861 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     4921 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
     4981 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     5041 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     5101 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
     5161 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     5221 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     5281 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     5341 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     5401 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     5461 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     5521 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     5581 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     5641 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     5701 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     5761 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     5821 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     5881 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     5941 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     6001 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     6061 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     6121 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     6181 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     6241 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     6301 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     6361 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     6421 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     6481 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     6541 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     6601 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     6661 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6721 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6781 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6841 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6901 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6961 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     7021 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     7081 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     7141 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     7201 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     7261 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
