LOCUS       sequence-375999-        8449 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 8449)
  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 8449)
  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: SFRS17A-1
FEATURES             Location/Qualifiers
     source          1..8449
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..1179
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          232..1170
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1127..1147
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1196..1215
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1196..1214
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1293..1317
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3406..3430)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3555..3596
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(4051..4068)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4057..4281
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4414..4433)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4565..4612
                     /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             4620..5642
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4620..5642
                     /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    5772..5893
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5809..5828)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5863..5882
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5942..5958)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5942..5958)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5955..5977)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5966..5982
                     /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(5990..5996,5997..6014,6015..6020))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6035..6056
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6173..6190)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6344..6932)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6424..6443)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7103..7894,7895..7963))
                     /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     7726..7745
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7964..8068)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8143..8162)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
       61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
      901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
      961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata
     1201 cgactcacta tagggagacc caagctggct aggtaagctt ggtaccgagc tcggatccac
     1261 tagtccagtg tggtggaatt ctgcagatat caacaagttt gtacaaaaaa gttggcacca
     1321 tggcagcggc taccatcgtg cacgacacgt ctgaggccgt ggagctctgc cctgcttacg
     1381 gcttgtacct gaagcccatc accaagatga ccatcagcgt ggcactcccg cagctgaagc
     1441 agccggggaa gtccatctcc aactgggagg tgatggagag gctgaagggc atggtgcaga
     1501 accaccagtt ctccacgctg cgtatttcca agagcaccat ggacttcatc cgcttcgagg
     1561 gggaggtgga gaacaagagc ctggtcaagt cttttctggc ctgcctggac ggcaagacca
     1621 tcaagctcag cggcttctcc gacatcctga aggtgcgcgc ggccgagttc aagatcgact
     1681 tccccacccg ccacgactgg gactccttct tccgcgacgc caaggacatg aacgagaccc
     1741 tgccggggga gcggccggac accatccacc tggaggggct gccctgcaag tggttcgccc
     1801 tgaaggagtc gggctccgag aagcccagcg aggacgtcct ggtcaaggtg tttgagaagt
     1861 tcggggagat ccggaatgtg gacatcccca tgctggaccc ctaccgggag gagatgacgg
     1921 gccgcaactt ccacaccttc agtttcgggg ggcacttgaa cttcgaggcc tatgtgcagt
     1981 accgtgagta catgggcttc atccaggcca tgagcgccct gcgcgggatg aaactcatgt
     2041 acaagggcga ggacggcaag gccgtggcct gcaacatcaa ggtttctttt gattcgacca
     2101 aacacctgag tgatgcctca attaagaagc ggcagctgga gaggcagaag cttcaggaac
     2161 tggagcagca aagagaagaa caaaagcgca gagagaagga agcggaggag aggcagcgag
     2221 cggaggaaag gaaacaaaag gagctggaag agctggagcg agagaggaaa agagaagaga
     2281 agcttcgcaa gagggagcag aagcagaggg accgtgagct gcgccggaat cagaagaagc
     2341 tggagaagct gcaggcggag gagcagaagc agctgcagga gaagatcaag ctggaggagc
     2401 gcaagctgct gctggcccag aggaacctgc agtccatccg gctcatcgcc gagctgctca
     2461 gcagagccaa ggctgtgaag ctacgggaac aggagcagaa ggaggagaag ctgaggctcc
     2521 agcagcagga ggagcggcgg cggctgcagg aggccgagct gcggcgcgtg gaggaggaga
     2581 aggagcgcgc gctgggcctg cagcggaaag agcgggagct gcgcgagcgg ctgctgagca
     2641 tcctgctgag caagaagccg gacgacagcc acacacacga cgagctgggc gtggcacacg
     2701 ccgacctgct gcagcccgtc ctggacatcc tgcagaccgt gtcgtccggc tgtgtgagcg
     2761 ccaccacgct gcaccccctc gggggccagc ccccggccgg tgcccccaag gagagcgcgg
     2821 cccacccaga ggccgacggc gctcccaaaa gcgtgaacgg gagcgtggcc gaggaggccc
     2881 catgcaagga ggttcagagc tcctgtcgtg tggtccccga ggatggctct ccagagaaga
     2941 ggtgcccggg cggcgtcctc tcctgcattc ctgacaacaa ccaacagccc aagggcatcc
     3001 ctgcctgcga gcagaatgtc tccagaaagg acacccggtc agaacaggac aagtgcaacc
     3061 gggagcccag caagggccgg ggccgggcca ccggagacgg gcttgctgac cggcacaagc
     3121 gggagaggag ccgggccagg cgggccagca gcagggagga cgggaggcca cgcaaggagc
     3181 ggcggcccca caagaagcac gcctacaagg atgacagccc ccgccggcgc agcacgagcc
     3241 cagaccacac ccggtcccgg aggtcccaca gcaaagacag gcaccggagg gagcggagcc
     3301 gggagcggag gggcagcgcc agcaggaagc acagccgcca ccgccgccga agcgagcggt
     3361 cgcgctcccg gtccccgagc aggcaccgca gtacctggaa caggttgcca actttcttgt
     3421 acaaagtggt tgatatccag cacagtggcg gccgctcgag tctagagggc ccgcggttcg
     3481 aaaccagcgg aggcggcgga agcgggggag gaggcagcgg cggaggcggc agcgggggcg
     3541 gcggaagcgc taccggtaag cctatcccta accctctcct cggtctcgat tctacgcgta
     3601 ccggtatggc ttctaacttt actcagttcg ttctcgtcga caatggcgga actggcgacg
     3661 tgactgtcgc cccaagcaac ttcgctaacg gggtcgctga atggatcagc tctaactcgc
     3721 gatcacaggc ttacaaagta acctgtagcg ttcgtcagag ctctgcgcag aatcgcaaat
     3781 acaccatcaa agtcgaggtg cctaaagtgg caacccagac tgttggtggt gaagagcttc
     3841 ctgtagccgg atggagatct tacttaaata tggaactaac cattccaatt ttcgccacga
     3901 attccgactg cgagcttatt gttaaggcaa tgcaaggtct cctaaaagat ggaaacccga
     3961 ttccctcggc catcgcggcc aactccggca tctacggagg tggaggtgga tctcgtacgg
     4021 gttagtaatg agtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat
     4081 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     4141 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     4201 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     4261 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt
     4321 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     4381 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     4441 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     4501 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     4561 cctagaagtt cctattccga agttcctatt ctctagaaag tataggaact tccttggcca
     4621 aaaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg
     4681 tctccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc ttcgatgtag
     4741 gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac aaagatcgtt
     4801 atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt gacattgggg
     4861 aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc acgttgcaag
     4921 acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc atggatgcga
     4981 tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg caaggaatcg
     5041 gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat gtgtatcact
     5101 ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc gatgagctga
     5161 tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca
     5221 acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc gaggcgatgt
     5281 tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta
     5341 tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga tcgccgcggc
     5401 tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg gttgacggca
     5461 atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga tccggagccg
     5521 ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc gatggctgtg
     5581 tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat
     5641 agcacgtact acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     5701 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     5761 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     5821 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     5881 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     5941 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     6001 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     6061 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     6121 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     6181 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     6241 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     6301 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     6361 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     6421 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     6481 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     6541 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     6601 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     6661 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     6721 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     6781 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     6841 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     6901 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     6961 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     7021 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     7081 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     7141 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     7201 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     7261 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     7321 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     7381 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     7441 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     7501 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     7561 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     7621 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     7681 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     7741 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     7801 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     7861 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     7921 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     7981 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     8041 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     8101 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
     8161 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
     8221 gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct
     8281 tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg
     8341 attattgact aggcttttgc aaaaagcttt gcaaagatgg ataaagtttt aaacagagag
     8401 gaatctttgc agctaatgga ccttctaggt cttgaaagga gtgggaatt
//
