LOCUS       sequence-376737-        8212 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For use in RBP tethering screen.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8212)
  AUTHORS   Luo EC, Nathanson JL, Tan FE, Schwartz JL, Schmok JC, Shankar A, 
            Markmiller S, Yee BA, Sathe S, Pratt GA, Scaletta DB, Ha Y, Hill DE,
            Aigner S, Yeo GW
  TITLE     Large-scale tethered function assays identify factors that regulate 
            mRNA stability and translation.
  JOURNAL   Nat Struct Mol Biol. 2020 Oct;27(10):989-1000. doi: 
            10.1038/s41594-020-0477-6. Epub 2020 Aug 17.
  PUBMED    32807991
REFERENCE   2  (bases 1 to 8212)
  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"; doi: 
            "10.1038/s41594-020-0477-6"; journalName: "Nat Struct Mol Biol"; 
            date: "2020-10"; volume: "27"; issue: "10"; pages: "989-1000"
COMMENT     Sequence Label: DDX5
FEATURES             Location/Qualifiers
     source          1..8212
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        121..1299
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          352..1290
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1247..1267
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1316..1335
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1316..1334
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1413..1437
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3289..3313)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3438..3479
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3934..3951)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3940..4164
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4297..4316)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4448..4495
                     /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             4503..5525
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4503..5525
                     /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    5655..5776
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5692..5711)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5746..5765
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5825..5841)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5825..5841)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5838..5860)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5849..5865
                     /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(5873..5879,5880..5897,5898..5903))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5918..5939
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6056..6073)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6227..6815)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6307..6326)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6986..7777,7778..7846))
                     /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     7609..7628
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7847..7951)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8026..8045)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgttgacatt gattattgac taggcttttg caaaaagctt tgcaaagatg gataaagttt
       61 taaacagaga ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt
      121 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
      181 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      241 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      301 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      361 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      421 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      481 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      541 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      601 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      661 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      721 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      781 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      841 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      901 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      961 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1021 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1081 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1141 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1201 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1261 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata
     1321 cgactcacta tagggagacc caagctggct aggtaagctt ggtaccgagc tcggatccac
     1381 tagtccagtg tggtggaatt ctgcagatat caacaagttt gtacaaaaaa gcaggctcca
     1441 ccatgtcggg ttattcgagt gaccgagacc gcggccggga ccgagggttt ggtgcacctc
     1501 gatttggagg aagtagggca gggcccttat ctggaaagaa gtttggaaac cctggggaga
     1561 aattagttaa aaagaagtgg aatcttgatg agctgcctaa atttgagaag aatttttatc
     1621 aagagcaccc tgatttggct aggcgcacag cacaagaggt ggaaacatac agaagaagca
     1681 aggaaattac agttagaggt cacaactgcc cgaagccagt tctaaatttt tatgaagcca
     1741 atttccctgc aaatgtcatg gatgttattg caagacagaa tttcactgaa cccactgcta
     1801 ttcaagctca gggatggcca gttgctctaa gtggattgga tatggttgga gtggcacaga
     1861 ctggatctgg gaaaacattg tcttatttgc ttcctgccat tgtccacatc aatcatcagc
     1921 cattcctaga gagaggcgat gggcctattt gtttggtgct ggcaccaact cgggaactgg
     1981 cccaacaggt gcagcaagta gctgctgaat attgtagagc atgtcgcttg aagtctactt
     2041 gtatctacgg tggtgctcct aagggaccac aaatacgtga tttggagaga ggtgtggaaa
     2101 tctgtattgc aacacctgga agactgattg actttttaga gtgtggaaaa accaatctga
     2161 gaagaacaac ctaccttgtc cttgatgaag cagatagaat gcttgatatg ggctttgaac
     2221 cccaaataag gaagattgtg gatcaaataa gacctgatag gcaaactcta atgtggagtg
     2281 cgacttggcc aaaagaagta agacagcttg ctgaagattt cctgaaagac tatattcata
     2341 taaacattgg tgcacttgaa ctgagtgcaa accacaacat tcttcagatt gtggatgtgt
     2401 gtcatgacgt agaaaaggat gaaaaactta ttcgtctaat ggaagagatc atgagtgaga
     2461 aggagaataa aaccattgtt tttgtggaaa ccaaaagaag atgtgatgag cttaccagaa
     2521 aaatgaggag agatgggtgg cctgccatgg gtatccatgg tgacaagagt caacaagagc
     2581 gtgactgggt tctaaatgaa ttcaaacatg gaaaagctcc tattctgatt gctacagatg
     2641 tggcctccag agggctagat gtggaagatg tgaaatttgt catcaattat gactacccta
     2701 actcctcaga ggattatatt catcgaattg gaagaactgc tcgcagtacc aaaacaggca
     2761 cagcatacac tttctttaca cctaataaca taaagcaagt gagcgacctt atctctgtgc
     2821 ttcgtgaagc taatcaagca attaatccca agttgcttca gttggtcgaa gacagaggtt
     2881 caggtcgttc caggggtaga ggaggcatga aggatgaccg tcgggacaga tactctgcgg
     2941 gcaaaagggg tggatttaat acctttagag acagggaaaa ttatgacaga ggttactcta
     3001 gcctgcttaa aagagatttt ggggcaaaaa ctcagaatgg tgtttacagt gctgcaaatt
     3061 acaccaatgg gagctttgga agtaattttg tgtctgctgg tatacagacc agttttagga
     3121 ctggtaatcc aacagggact taccagaatg gttatgatag cactcagcaa tacggaagta
     3181 atgttccaaa tatgcacaat ggtatgaacc aacaggcata tgcatatcct gctactgcag
     3241 ctgcacctat gattggttat ccaatgccaa caggatattc ccaattggac ccagctttct
     3301 tgtacaaagt ggttgatatc cagcacagtg gcggccgctc gagtctagag ggcccgcggt
     3361 tcgaaaccag cggaggcggc ggaagcgggg gaggaggcag cggcggaggc ggcagcgggg
     3421 gcggcggaag cgctaccggt aagcctatcc ctaaccctct cctcggtctc gattctacgc
     3481 gtaccggtat ggcttctaac tttactcagt tcgttctcgt cgacaatggc ggaactggcg
     3541 acgtgactgt cgccccaagc aacttcgcta acggggtcgc tgaatggatc agctctaact
     3601 cgcgatcaca ggcttacaaa gtaacctgta gcgttcgtca gagctctgcg cagaatcgca
     3661 aatacaccat caaagtcgag gtgcctaaag tggcaaccca gactgttggt ggtgaagagc
     3721 ttcctgtagc cggatggaga tcttacttaa atatggaact aaccattcca attttcgcca
     3781 cgaattccga ctgcgagctt attgttaagg caatgcaagg tctcctaaaa gatggaaacc
     3841 cgattccctc ggccatcgcg gccaactccg gcatctacgg aggtggaggt ggatctcgta
     3901 cgggttagta atgagtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     3961 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     4021 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     4081 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     4141 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     4201 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     4261 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     4321 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     4381 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     4441 gtacctagaa gttcctattc cgaagttcct attctctaga aagtatagga acttccttgg
     4501 ccaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca
     4561 gcgtctccga cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg
     4621 taggagggcg tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc
     4681 gttatgttta tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg
     4741 gggaattcag cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc
     4801 aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg
     4861 cgatcgctgc ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa
     4921 tcggtcaata cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc
     4981 actggcaaac tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc
     5041 tgatgctttg ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct
     5101 ccaacaatgt cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga
     5161 tgttcgggga ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt
     5221 gtatggagca gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc
     5281 ggctccgggc gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg
     5341 gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag
     5401 ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct
     5461 gtgtagaagt actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg
     5521 aatagcacgt actacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     5581 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     5641 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     5701 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     5761 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
     5821 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     5881 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     5941 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     6001 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     6061 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     6121 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     6181 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     6241 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     6301 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     6361 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     6421 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     6481 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     6541 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     6601 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     6661 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     6721 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     6781 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     6841 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     6901 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     6961 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     7021 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     7081 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     7141 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     7201 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     7261 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     7321 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     7381 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     7441 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     7501 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     7561 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     7621 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     7681 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     7741 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     7801 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     7861 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     7921 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     7981 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
     8041 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
     8101 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
     8161 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cg
//
