LOCUS       sequence-376042-        6670 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 6670)
  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 6670)
  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: CSTF3
FEATURES             Location/Qualifiers
     source          1..6670
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        67..1245
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          298..1236
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1193..1213
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1262..1281
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1262..1280
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1359..1383
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1693..1717)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1842..1883
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2338..2355)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2344..2568
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2701..2720)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2852..2899
                     /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             2907..3929
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2907..3929
                     /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    4059..4180
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4096..4115)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4150..4169
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4229..4245)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4229..4245)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4242..4264)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4253..4269
                     /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(4277..4283,4284..4301,4302..4307))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4322..4343
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4460..4477)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4631..5219)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4711..4730)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5390..6181,6182..6250))
                     /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     6013..6032
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6251..6355)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6430..6449)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aagttttaaa cagagaggaa tctttgcagc taatggacct tctaggtctt gaaaggagtg
       61 ggaattggct ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag
      121 ttggggggag gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg
      181 gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata
      241 agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacaggta
      301 agtgccgtgt gtggttcccg cgggcctggc ctctttacgg gttatggccc ttgcgtgcct
      361 tgaattactt ccacctggct gcagtacgtg attcttgatc ccgagcttcg ggttggaagt
      421 gggtgggaga gttcgaggcc ttgcgcttaa ggagcccctt cgcctcgtgc ttgagttgag
      481 gcctggcctg ggcgctgggg ccgccgcgtg cgaatctggt ggcaccttcg cgcctgtctc
      541 gctgctttcg ataagtctct agccatttaa aatttttgat gacctgctgc gacgcttttt
      601 ttctggcaag atagtcttgt aaatgcgggc caagatctgc acactggtat ttcggttttt
      661 ggggccgcgg gcggcgacgg ggcccgtgcg tcccagcgca catgttcggc gaggcggggc
      721 ctgcgagcgc ggccaccgag aatcggacgg gggtagtctc aagctggccg gcctgctctg
      781 gtgcctggcc tcgcgccgcc gtgtatcgcc ccgccctggg cggcaaggct ggcccggtcg
      841 gcaccagttg cgtgagcgga aagatggccg cttcccggcc ctgctgcagg gagctcaaaa
      901 tggaggacgc ggcgctcggg agagcgggcg ggtgagtcac ccacacaaag gaaaagggcc
      961 tttccgtcct cagccgtcgc ttcatgtgac tccacggagt accgggcgcc gtccaggcac
     1021 ctcgattagt tctcgagctt ttggagtacg tcgtctttag gttgggggga ggggttttat
     1081 gcgatggagt ttccccacac tgagtgggtg gagactgaag ttaggccagc ttggcacttg
     1141 atgtaattct ccttggaatt tgcccttttt gagtttggat cttggttcat tctcaagcct
     1201 cagacagtgg ttcaaagttt ttttcttcca tttcaggtgt cgtgaggaat tagcttggta
     1261 ctaatacgac tcactatagg gagacccaag ctggctaggt aagcttggta ccgagctcgg
     1321 atccactagt ccagtgtggt ggaattctgc agatatcaac aagtttgtac aaaaaagttg
     1381 gcatgtcagg agacggagcc acggagcagg cagctgagta tgtcccagag aaggtgaaga
     1441 aagcggaaaa gaaattagaa gagaatccat atgaccttga tgcttggagc attctcattc
     1501 gagaggcaca gaatcaacct atagacaaag cacggaagac ttatgaacgc cttgttgccc
     1561 agttccccag ttctggcaga ttctggaaac tgtacattga agcagaggtt actattttat
     1621 tttatttttt cttatatcag tattgcagca ttcactgtag tgatagaaaa caagttagga
     1681 acatagccaa ttacccaact ttcttgtaca aagtggttga tatccagcac agtggcggcc
     1741 gctcgagtct agagggcccg cggttcgaaa ccagcggagg cggcggaagc gggggaggag
     1801 gcagcggcgg aggcggcagc gggggcggcg gaagcgctac cggtaagcct atccctaacc
     1861 ctctcctcgg tctcgattct acgcgtaccg gtatggcttc taactttact cagttcgttc
     1921 tcgtcgacaa tggcggaact ggcgacgtga ctgtcgcccc aagcaacttc gctaacgggg
     1981 tcgctgaatg gatcagctct aactcgcgat cacaggctta caaagtaacc tgtagcgttc
     2041 gtcagagctc tgcgcagaat cgcaaataca ccatcaaagt cgaggtgcct aaagtggcaa
     2101 cccagactgt tggtggtgaa gagcttcctg tagccggatg gagatcttac ttaaatatgg
     2161 aactaaccat tccaattttc gccacgaatt ccgactgcga gcttattgtt aaggcaatgc
     2221 aaggtctcct aaaagatgga aacccgattc cctcggccat cgcggccaac tccggcatct
     2281 acggaggtgg aggtggatct cgtacgggtt agtaatgagt ttaaacccgc tgatcagcct
     2341 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     2401 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     2461 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     2521 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
     2581 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
     2641 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     2701 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     2761 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     2821 aacttgatta gggtgatggt tcacgtacct agaagttcct attccgaagt tcctattctc
     2881 tagaaagtat aggaacttcc ttggccaaaa agcctgaact caccgcgacg tctgtcgaga
     2941 agtttctgat cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag
     3001 aatctcgtgc tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct
     3061 gcgccgatgg tttctacaaa gatcgttatg tttatcggca ctttgcatcg gccgcgctcc
     3121 cgattccgga agtgcttgac attggggaat tcagcgagag cctgacctat tgcatctccc
     3181 gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc
     3241 agccggtcgc ggaggccatg gatgcgatcg ctgcggccga tcttagccag acgagcgggt
     3301 tcggcccatt cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg
     3361 cgattgctga tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt
     3421 ccgtcgcgca ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc
     3481 acctcgtgca cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag
     3541 cggtcattga ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct
     3601 tcttctggag gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc
     3661 atccggagct tgcaggatcg ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc
     3721 aactctatca gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat
     3781 gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa
     3841 gcgcggccgt ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc
     3901 ccagcactcg tccgagggca aaggaatagc acgtactacg agatttcgat tccaccgccg
     3961 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     4021 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     4081 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4141 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     4201 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     4261 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     4321 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     4381 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     4441 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     4501 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     4561 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     4621 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     4681 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     4741 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     4801 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4861 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4921 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4981 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5041 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5101 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5161 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5221 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     5281 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     5341 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     5401 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     5461 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     5521 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     5581 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     5641 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     5701 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     5761 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     5821 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     5881 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     5941 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6001 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6061 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6121 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6181 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6241 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     6301 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     6361 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg
     6421 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
     6481 tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt
     6541 gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
     6601 acgggccaga tatacgcgtt gacattgatt attgactagg cttttgcaaa aagctttgca
     6661 aagatggata
//
