LOCUS       sequence-392974-        8419 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of human FXR2 protein in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8419)
  AUTHORS   Edwards M, Xu M, Joseph S
  TITLE     A simple procedure for bacterial expression and purification of the 
            fragile X protein family.
  JOURNAL   Sci Rep. 2020 Sep 28;10(1):15858. doi: 10.1038/s41598-020-72984-7.
  PUBMED    32985615
REFERENCE   2  (bases 1 to 8419)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Rep."; 
            date: "2020-09-28"; volume: "10(1):15858. doi"; pages: " 
            10.1038/s41598-020-72984-7"
COMMENT     Sequence Label: His6_MBP_FXR2P_P474S_P493N
FEATURES             Location/Qualifiers
     source          1..8419
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      5..593
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     494..513
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     747..764
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    779..918
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(844..863)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1004..1026
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1020..1211)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2020..3102)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLSTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     3064..3083
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3103..3180)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3386..3405
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3493..3512
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3493..3511
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3512..3536
                     /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)."
     RBS             3551..3573
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3584..3601
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3626..4723
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="KIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      4580..4589
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     4697..4720
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             4739..4762
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6848..6865
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6918..6936)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6932..6979
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(7103..7122)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        7289..7392
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7393..7461,7462..8253)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7611..7630)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
       61 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
      121 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
      181 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
      241 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
      301 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
      361 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
      421 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
      481 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
      541 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
      601 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
      661 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
      721 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
      781 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct
      841 cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt
      901 gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
      961 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     1021 cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg
     1081 tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc
     1141 tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc
     1201 tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg
     1261 atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg
     1321 gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact
     1381 cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag
     1441 catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga
     1501 ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg
     1561 cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg
     1621 caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg
     1681 gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg
     1741 acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc
     1801 gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt
     1861 cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc
     1921 gcccaccgga aggagctgac tgggttgaag gctttcaagg gcatcggtcg agatcccggt
     1981 gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg
     2041 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     2101 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
     2161 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
     2221 gcgaaaatcc tgtttgatgg tggataacgg cgggatataa catgagctgt cttcggtatc
     2281 gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg
     2341 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
     2401 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
     2461 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
     2521 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
     2581 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
     2641 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
     2701 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
     2761 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
     2821 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
     2881 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
     2941 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
     3001 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
     3061 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
     3121 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
     3181 cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
     3241 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
     3301 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
     3361 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
     3421 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atcgatctcg
     3481 atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac aattcccctc
     3541 tagaaataat tttgtttaac tttaagaagg agatatacat atgcaccatc atcatcatca
     3601 ttcttctggt gtagatctgg gtaccaaaat cgaagaaggt aaactggtaa tctggattaa
     3661 cggcgataaa ggctataacg gtctcgctga agtcggtaag aaattcgaga aagataccgg
     3721 aattaaagtc accgttgagc atccggataa actggaagag aaattcccac aggttgcggc
     3781 aactggcgat ggccctgaca ttatcttctg ggcacacgac cgctttggtg gctacgctca
     3841 atctggcctg ttggctgaaa tcaccccgga caaagcgttc caggacaagc tgtatccgtt
     3901 tacctgggat gccgtacgtt acaacggcaa gctgattgct tacccgatcg ctgttgaagc
     3961 gttatcgctg atttataaca aagatctgct gccgaacccg ccaaaaacct gggaagagat
     4021 cccggcgctg gataaagaac tgaaagcgaa aggtaagagc gcgctgatgt tcaacctgca
     4081 agaaccgtac ttcacctggc cgctgattgc tgctgacggg ggttatgcgt tcaagtatga
     4141 aaacggcaag tacgacatta aagacgtggg cgtggataac gctggcgcga aagcgggtct
     4201 gaccttcctg gttgacctga ttaaaaacaa acacatgaat gcagacaccg attactccat
     4261 cgcagaagct gcctttaata aaggcgaaac agcgatgacc atcaacggcc cgtgggcatg
     4321 gtccaacatc gacaccagca aagtgaatta tggtgtaacg gtactgccga ccttcaaggg
     4381 tcaaccatcc aaaccgttcg ttggcgtgct gagcgcaggt attaacgccg ccagtccgaa
     4441 caaagagctg gcaaaagagt tcctcgaaaa ctatctgctg actgatgaag gtctggaagc
     4501 ggttaataaa gacaaaccgc tgggtgccgt agcgctgaag tcttacgagg aagagttggc
     4561 gaaagatcca cgtattgccg ccaccatgga aaacgcccag aaaggtgaaa tcatgccgaa
     4621 catcccgcag atgtccgctt tctggtatgc cgtgcgtact gcggtgatca acgccgccag
     4681 cggtcgtcag actgtcgatg aagccctgaa agacgcgcag actggtggtg gtggtagcct
     4741 ggaagttctg ttccagggtc cgatgggcgg tctggcgagc ggtggcgatg ttgagccggg
     4801 tctgccggtg gaagtgcgtg gtagcaatgg tgcgttttac aagggttttg ttaaagacgt
     4861 gcacgaggac agcgttacca tcttctttga aaacaactgg caaagcgaac gtcagatccc
     4921 gtttggtgac gtgcgtctgc cgccgccggc ggactataac aaggaaatca ccgaaggcga
     4981 tgaggtggaa gtttatagcc gtgcgaacga gcaagaaccg tgcggttggt ggctggcgcg
     5041 tgttcgtatg atgaagggtg acttctacgt gattgaatat gcggcgtgcg atgcgaccta
     5101 caacgagatc gttaccctgg aacgtctgcg tccggttaac ccgaacccgc tggcgaccaa
     5161 aggtagcttc tttaaggtga cgatggcggt gccggaggat ctgcgtgaag cgtgcagcaa
     5221 cgaaaacgtg cacaaggagt tcaagaaggc gctgggcgcg aactgcatct tcctgaacat
     5281 caccaacagc gagctgttca tcctgagcac caccgaggcg ccggttaaac gtgcgagcct
     5341 gctgggtgac atgcacttcc gtagcctgcg taccaagctg ctgctgatga gccgtaacga
     5401 agaggcgacc aaacacctgg agaccagcaa gcaactggcg gcggcgtttc aggaagagtt
     5461 caccgtgcgt gaggacctga tgggtctggc gattggcacc cacggtgcga acattcaaca
     5521 ggcgcgtaaa gttccgggcg ttaccgcgat cgagctgggt gaagagacct gcaccttccg
     5581 tatctacggt gaaaccccgg aggcgtgccg tcaagcgcgt agctacctgg agtttagcga
     5641 agatagcgtg caagttccgc gtaacctggt tggcaaagtg atcggcaaga acggtaaagt
     5701 gattcaggag atcgtggaca aaagcggtgt ggttcgtgtg cgtgttgagg gtgacaacga
     5761 taagaaaaac ccgcgtgaag agggtatggt gccgtttatt ttcgtgggca cccgtgagaa
     5821 catcagcaac gcgcaagcgc tgctggagta tcacctgagc tacctgcaag aggtggaaca
     5881 gctgcgtctg gagcgtctgc agattgacga gcaactgcgt caaattggcc tgggttttcg
     5941 tccgccgggt agcggccgtg gtagcggcgg tagcgacaag gcgggttata gcaccgacga
     6001 aagcagcagc agcagcctgc acgcgacccg tacctacggt ggcagctatg gtggccgtgg
     6061 tcgtggccgt cgtaccggtg gcccggcgta tggtccgagc agcgatgtga gcaccgcgag
     6121 cgaaaccgag agcgaaaagc gtgaagagcc gaaccgtgcg ggtccgggtg accgtgatcc
     6181 gagcacccgt ggtgaagaaa gccgtcgtcg tccgaccggt ggtcgtggcc gtggtccgaa
     6241 cccggcgccg cgtccgacca gccgttacaa cagcagcagc attagcagcg tgctgaaaga
     6301 cccggatagc aacccgtata gcctgctgga caccagcgaa ccggaaccgc cggtggatag
     6361 cgagccgggc gaaccgccgc cggcgagcgc gcgtcgtcgt cgtagccgtc gtcgtcgtac
     6421 cgatgaagac cgtaccgtga tggatggcgg cctggaaagc gacggtccga acatgaccga
     6481 gaacggtctg gaggacgaaa gccgtccgca acgtcgtaac cgtagccgtc gtcgtcgtaa
     6541 ccgtggtaac cgtaccgatg gcagcatcag cggtgatcgt caaccggtta ccgtggcgga
     6601 ttacatcagc cgtgcggaga gccaaagccg tcagcgtccg ccgctggagc gtaccaaacc
     6661 gagcgaggat agcctgagcg gccagaaagg tgacagcgtg agcaaactgc cgaagggtcc
     6721 gagcgagaat ggtgaactga gcgcgccgct ggaactgggt agcatggtga atggtgtgag
     6781 ctaacattgg aagtggataa cggatccgaa ttcgagctcc gtcgacaagc ttgcggccgc
     6841 actcgagcac caccaccacc accactgaga tccggctgct aacaaagccc gaaaggaagc
     6901 tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg
     6961 ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg
     7021 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     7081 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     7141 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     7201 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacatta acgcttacaa
     7261 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     7321 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatttga
     7381 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     7441 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     7501 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     7561 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     7621 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     7681 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     7741 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     7801 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     7861 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     7921 gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     7981 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     8041 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     8101 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     8161 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     8221 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     8281 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     8341 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     8401 gtagaaaaga tcaaaggat
//
