LOCUS       sequence-448451-        7777 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Express mEGFP-fusion protein of FMR1 isoform 1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7777)
  AUTHORS   Chen X, Fansler MM, Janjos U, Ule J, Mayr C
  TITLE     The FXR1 network acts as signaling scaffold for actomyosin 
            remodeling.
  JOURNAL   Cell. 5 August 2024. doi: 10.1016/j.cell.2024.07.015.
  PUBMED    37961296
REFERENCE   2  (bases 1 to 7777)
  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"; journalName: "Cell. 5 
            August 2024. doi: 10.1016/j.cell.2024.07.015."
COMMENT     Sequence Label: pcDNA3puro-mGFP-FMR1 isoform1-sh1Resistant
FEATURES             Location/Qualifiers
     source          1..7777
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(905..907,908..910,911..1621)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(950..971)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1211..1230)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1558..1579
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3617..3634)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3623..3847
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3893..4321
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3980..3999)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4190..4211
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4330..4350)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4335..4664
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4515..4650
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4577..4596
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4677..5276
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4677..4696)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5173..5193
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5450..5571
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5487..5506)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5541..5560
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5620..5636)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5620..5636)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5633..5655)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5644..5660
                     /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(5668..5674,5675..5692,5693..5698))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5713..5734
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5851..5868)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6022..6610)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6102..6121)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6781..7572,7573..7641))
                     /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     7404..7423
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7642..7746)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 aaccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
      961 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     1021 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     1081 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     1141 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     1201 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1261 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1321 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     1381 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1441 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1501 ccactacctg agcacccagt ccaagctgag caaagacccc aacgagaagc gcgatcacat
     1561 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1621 aaaagcaggc tccaccatgg aggagctggt ggtggaagtg cggggctcca atggcgcttt
     1681 ctacaaggca tttgtaaagg atgttcatga agattcaata acagttgcat ttgaaaacaa
     1741 ctggcagcct gataggcaga ttccatttca tgatgtcaga ttcccacctc ctgtaggtta
     1801 taataaagat ataaatgaaa gtgatgaagt tgaggtgtat tccagagcaa atgaaaaaga
     1861 gccttgctgt tggtggttag ctaaagtgag gatgataaag ggtgagtttt atgtgataga
     1921 atatgcagca tgtgatgcaa cttacaatga aattgtcaca attgaacgtc taagatctgt
     1981 taatcccaac aaacctgcca caaaagatac tttccataag atcaagctgg atgtgccaga
     2041 agacttacgg caaatgtgtg ccaaagaggc ggcacataag gattttaaaa aggcagttgg
     2101 tgccttttct gtaacttatg atccagaaaa ttatcagctt gtcattttgt ccatcaatga
     2161 agtcacctca aagcgagcac atatgctgat tgacatgcac tttcggagtc tgcgcactaa
     2221 gttgtctctg ataatgagaa atgaagaagc tagtaagcag ctggagagtt caaggcagct
     2281 tgcctcgaga tttcatgaac agtttatcgt aagagaagat ctgatgggtc tagctattgg
     2341 tactcatggt gctaatattc agcaagctag aaaagtacct ggggtcactg ctattgatct
     2401 agatgaagat acctgcacat ttcatattta tggagaggat caggatgcag tgaaaaaagc
     2461 tagaagcttt ctcgaatttg ctgaagatgt aatacaagtt ccaaggaact tagtaggcaa
     2521 agtaatagga aaaaatggaa agctgattca ggagattgtg gacaagtcag gagttgtgag
     2581 ggtgaggatt gaggctgaaa atgagaaaaa tgttccacaa gaagaggaaa ttatgccacc
     2641 aaattccctt ccttccaata attcaagggt tggacctaat gccccagaag aaaaaaaaca
     2701 tttagatata aaggaaaaca gcacccattt ttctcaacct aacagtacaa aagtccagag
     2761 ggtgttagtg gcttcatcag ttgtagcagg ggaatcccag aaacctgaac tcaaggcttg
     2821 gcagggtatg gtaccatttg tttttgtggg aacaaaggac agcatcgcta atgccactgt
     2881 tcttttggat tatcacctga actatttaaa ggaagtagac cagttgagac tcgagcggct
     2941 acagattgat gagcagttgc gacagattgg agctagttct agaccaccac caaatcgtac
     3001 agataaggaa aaaagctatg tgactgatga tggtcaagga atgggtcgag gtagtagacc
     3061 ttacagaaat agggggcacg gcagacgcgg tcctggatat acttcaggaa ctaattctga
     3121 agcatcaaat gcttctgaaa cagaatctga ccacagagac gaactcagtg attggtcatt
     3181 agctccaaca gaggaagaga gggagagctt cctgcgcaga ggagacggac ggcggcgtgg
     3241 agggggagga agaggacaag gaggaagagg acgtggagga ggcttcaaag gaaacgacga
     3301 tcactcccga acagataatc gtccacgtaa tccaagagag gctaaaggaa gaacaacaga
     3361 tggatccctt cagatcagag ttgactgcaa taatgaaagg agtgtccaca ctaaaacatt
     3421 acagaatacc tccagtgaag gtagtcggct gcgcacgggt aaagatcgta accagaagaa
     3481 agagaagcca gacagcgtgg atggtcagca accactcgtg aatggagtac cctaaacgaa
     3541 ttcgcggccg cactcgagat tccagcacag tggcggccgc tcgagtctag agggcccgtt
     3601 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     3661 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     3721 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     3781 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     3841 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     3901 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     3961 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4021 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4081 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     4141 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     4201 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4261 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4321 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     4381 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     4441 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     4501 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     4561 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     4621 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccaaccatga
     4681 ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca
     4741 ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc
     4801 acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg
     4861 gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg
     4921 tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc
     4981 ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg
     5041 cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg
     5101 ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga
     5161 cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg
     5221 tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgagcgg
     5281 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     5341 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     5401 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     5461 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5521 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     5581 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     5641 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     5701 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     5761 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5821 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5881 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5941 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     6001 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     6061 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6121 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6181 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     6241 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6301 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6361 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6421 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     6481 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6541 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6601 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6661 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6721 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6781 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6841 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6901 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6961 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     7021 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     7081 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7141 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7201 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7261 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7321 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7381 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7441 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7501 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7561 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7621 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7681 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7741 tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
