LOCUS       sequence-448475-        7682 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Express mEGFP-fusion protein of FXR1 isoform a (I304N).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7682)
  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 7682)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 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-FXR1a-I304N
FEATURES             Location/Qualifiers
     source          1..7682
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      76..504
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(163..182)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     373..394
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(513..533)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        518..847
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      698..833
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     760..779
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             860..1459
                     /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(860..879)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1356..1376
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1633..1754
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1670..1689)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1724..1743
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1803..1819)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1803..1819)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1816..1838)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1827..1843
                     /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(1851..1857,1858..1875,1876..1881))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1896..1917
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2034..2051)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2205..2793)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2285..2304)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2964..3755,3756..3824))
                     /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     3587..3606
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3825..3929)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4004..4023)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4195..4574
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4575..4778
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4729..4749
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4823..4842
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4823..4841
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(4865..4867,4868..4870,4871..5581)
                     /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(4910..4931)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5171..5190)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5518..5539
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(7482..7499)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    join(7488..7682,1..30)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
       61 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
      121 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
      181 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
      241 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
      301 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
      361 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
      421 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
      481 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
      541 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
      601 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
      661 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
      721 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
      781 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
      841 ttgcaaaaag ctcccaacca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga
      901 cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg
      961 ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct
     1021 cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc
     1081 ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg
     1141 catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc
     1201 gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca
     1261 ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc
     1321 cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct
     1381 cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac
     1441 ccgcaagccc ggtgcctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     1501 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     1561 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     1621 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     1681 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     1741 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     1801 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     1861 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     1921 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     1981 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     2041 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     2101 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     2161 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2221 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2281 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2341 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2401 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2461 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2521 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     2581 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2641 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2701 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2761 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2821 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     2881 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     2941 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     3001 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     3061 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     3121 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     3181 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     3241 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     3301 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     3361 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     3421 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     3481 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     3541 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     3601 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3661 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3721 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     3781 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     3841 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     3901 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     3961 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
     4021 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
     4081 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
     4141 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
     4201 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     4261 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     4321 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     4381 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     4441 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     4501 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     4561 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
     4621 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
     4681 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
     4741 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
     4801 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
     4861 aaccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     4921 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     4981 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     5041 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     5101 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     5161 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     5221 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     5281 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     5341 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     5401 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     5461 ccactacctg agcacccagt ccaagctgag caaagacccc aacgagaagc gcgatcacat
     5521 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     5581 ggcggagctg acggtggagg ttcgcggctc taacggggct ttctacaagg gatttatcaa
     5641 agatgttcat gaagactccc ttacagttgt ttttgaaaat aattggcaac cagaacgcca
     5701 ggttccattt aatgaagtta gattaccacc accacctgat ataaaaaaag aaattagtga
     5761 aggagatgaa gtagaggtat attcaagagc aaatgaccaa gagccatgtg ggtggtggtt
     5821 ggctaaagtt cggatgatga aaggagaatt ttatgtcatt gaatatgctg cttgtgacgc
     5881 tacttacaat gaaatagtca catttgaacg acttcggcct gtcaatcaaa ataaaactgt
     5941 caaaaaaaat accttcttta aatgcacagt ggatgttcct gagcatttga gagaggcgtg
     6001 tgctaatgaa aatgcacata aagattttaa gaaagcagta ggagcatgca gaatttttta
     6061 ccatccagaa acaacacagc taatgatact gtctgccagt gaagcaactg tgaagagagt
     6121 aaacatctta agtgacatgc atttgcgaag tattcgtacg aagttgatgc ttatgtccag
     6181 aaatgaagag gccactaagc atttagaatg cacaaaacaa cttgcagcag cttttcatga
     6241 ggaatttgtt gtgagagaag atttaatggg cctggcaata ggaacacatg gtagtaacat
     6301 ccagcaagct aggaaggttc ctggagttac cgccattgag ctagatgaag atactggaac
     6361 attcagaatc tacggagaga gtgctgatgc tgtaaaaaag gctagaggtt tcttggaatt
     6421 tgtggaggat tttattcagg ttcctaggaa tctcgttgga aaagtaattg gaaaaaatgg
     6481 caaagttaat caagaaatag tggacaaatc tggtgtggtt cgagtgagaa ttgaagggga
     6541 caatgaaaat aaattaccca gagaagacgg tatggttcca tttgtatttg ttggcactaa
     6601 agaaagcatt ggaaatgtgc aggttcttct agagtatcat attgcctatc taaaggaagt
     6661 agaacagcta agaatggaac gcctacagat tgatgaacag ctgcgacaga ttggttctag
     6721 gtcttatagc ggaagaggca gaggtcgtcg gggacctaat tacacctccg gttatggtac
     6781 aaattctgag ctgtctaacc cctctgaaac ggaatctgag cgtaaagacg agctgagtga
     6841 ttggtcattg gcaggagaag atgatcgaga cagccgacat cagcgtgaca gcaggagacg
     6901 cccaggagga agaggcagaa gtgtttcagg gggtcgaggt cgtggtggac cacgtggtgg
     6961 caaatcctcc atcagttctg tgctcaaaga tccagacagc aatccataca gcttacttga
     7021 taatacagaa tcagatcaga ctgcagacac tgatgcatcc gaaagccacc attccactaa
     7081 ccgtcgtagg cggtctcgta gacgaaggac tgatgaagat gctgttctga tggatggaat
     7141 gactgaatct gatacagctt cagttaatga aaatgggcta gtcacagttg cagattatat
     7201 ttctagagct gagtctcaga gcagacaaag aaacctccca agggaaactt tggctaaaaa
     7261 caagaaagaa atggcaaaag atgtgattga agagcatggt ccttcagaaa aggcaataaa
     7321 cggcccaact agtgcttctg gcgatgacat ttctaagcta cagcgtactc caggagaaga
     7381 aaagattaat accttaaaag aagaaaacac tcaagaagca gcagtcctga atggtgtttc
     7441 ataactcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag
     7501 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     7561 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     7621 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     7681 ca
//
