LOCUS       sequence-377075-        8341 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 8341)
  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 8341)
  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: THOC1
FEATURES             Location/Qualifiers
     source          1..8341
                     /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(3298..3322)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3447..3488
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3943..3960)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3949..4173
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4306..4325)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4457..4504
                     /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             4512..5534
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDYICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    5664..5785
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5701..5720)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5755..5774
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5834..5850)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5834..5850)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5847..5869)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5858..5874
                     /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(5882..5888,5889..5906,5907..5912))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5927..5948
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6065..6082)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6236..6824)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6316..6335)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6995..7786,7787..7855))
                     /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     7618..7637
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7856..7960)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8035..8054)
                     /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 gcaggctcca
     1321 ccatgtctcc gacgccgccg ctcttcagtt tgcccgaagc gcggacgcgg tttacgaagt
     1381 ctaccagaga ggccttgaac aacaaaaaca tcaagccatt gttaagtacc ttcagccagg
     1441 tacctggcag tgaaaatgaa aaaaaatgta cccttgacca agctttcaga ggtattctag
     1501 aagaagaaat tataaatcat tcatcatgtg aaaacgtttt agctattatt tctcttgcta
     1561 ttgggggagt aactgaaggt atttgtaccg catctacacc ttttgtattg ttgggagatg
     1621 ttttggattg tcttcctttg gatcagtgtg acacaatatt cacttttgtg gaaaaaaatg
     1681 ttgctacttg gaaatcaaat acattctatt ctgctgggaa aaattactta ctacgtatgt
     1741 gcaatgatct cctaagaaga ttgtctaaat cccagaatac agtcttctgt ggacggattc
     1801 agctcttttt ggccaggctt ttccctctgt ctgagaaatc aggtcttaac ttgcagagtc
     1861 agtttaatct ggaaaatgtc actgttttca atacaaatga gcaggaaagc accctgggtc
     1921 agaagcacac tgaagataga gaagaaggaa tggatgtaga agaaggcgaa atgggagatg
     1981 aggaagctcc aacaacgtgc tctattccaa ttgattacaa cctgtatcga aaattctggt
     2041 cacttcagga ttacttcagg aaccctgtgc aatgctatga gaagatttca tggaaaactt
     2101 ttctcaagta ttctgaagaa gttttagctg tttttaagag ttataaatta gatgatactc
     2161 aggcctcaag aaaaaagatg gaagaattga aaacaggagg agaacatgta tattttgcaa
     2221 aatttttaac aagtgaaaag ctgatggatt tacaactgag tgacagtaac tttcgtcgac
     2281 acatcctgtt gcagtatctc attttattcc aatatctcaa ggggcaggtc aaattcaaaa
     2341 gttcaaacta tgttttaact gatgagcaat cactttggat tgaagatact acaaaatcag
     2401 tttatcaact actatctgaa aacccccccg atggagaaag attttcaaag atggtagagc
     2461 atatattaaa cactgaagaa aactggaact cgtggaaaaa tgaaggttgc ccaagttttg
     2521 tgaaagaaag aacatcagat accaaaccta cgagaataat tcggaagaga acagcacccg
     2581 aggacttcct agggaaagga cccaccaaaa aaattctgat gggaaatgag gagttaacaa
     2641 ggctttggaa tctttgccct gataatatgg aagcctgtaa atcagagaca agggaacaca
     2701 tgcccacttt ggaggaattc tttgaagaag ccattgaaca ggcagaccct gaaaatatgg
     2761 tggaaaatga atataaggct gtgaacaatt caaattatgg ttggagagcc ctgagactat
     2821 tagcacggag aagccctcac ttcttccagc caaccaacca gcagtttaaa agtttaccag
     2881 aatatcttga aaatatggta ataaagctag ccaaggaatt accgcctcct tctgaagaaa
     2941 taaaaacagg tgaggatgaa gatgaggaag ataatgatgc tctactgaag gaaaatgaaa
     3001 gtcctgatgt tcggcgagac aaacctgtaa caggagaaca aatagaggta tttgccaaca
     3061 agctgggtga acaatggaag attctggctc cctacttgga aatgaaagac tcagaaatta
     3121 ggcagattga gtgtgacagt gaagacatga agatgagagc taagcagctc ctggttgcct
     3181 ggcaagatca agagggagtt catgcaacac ctgagaatct gattaatgca ctgaataagt
     3241 ctggattaag tgaccttgca gaaagtctaa ctaatgacaa tgagacaaat agtttggacc
     3301 cagctttctt gtacaaagtg gttgatatcc agcacagtgg cggccgctcg agtctagagg
     3361 gcccgcggtt cgaaaccagc ggaggcggcg gaagcggggg aggaggcagc ggcggaggcg
     3421 gcagcggggg cggcggaagc gctaccggta agcctatccc taaccctctc ctcggtctcg
     3481 attctacgcg taccggtatg gcttctaact ttactcagtt cgttctcgtc gacaatggcg
     3541 gaactggcga cgtgactgtc gccccaagca acttcgctaa cggggtcgct gaatggatca
     3601 gctctaactc gcgatcacag gcttacaaag taacctgtag cgttcgtcag agctctgcgc
     3661 agaatcgcaa atacaccatc aaagtcgagg tgcctaaagt ggcaacccag actgttggtg
     3721 gtgaagagct tcctgtagcc ggatggagat cttacttaaa tatggaacta accattccaa
     3781 ttttcgccac gaattccgac tgcgagctta ttgttaaggc aatgcaaggt ctcctaaaag
     3841 atggaaaccc gattccctcg gccatcgcgg ccaactccgg catctacgga ggtggaggtg
     3901 gatctcgtac gggttagtaa tgagtttaaa cccgctgatc agcctcgact gtgccttcta
     3961 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     4021 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     4081 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     4141 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     4201 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     4261 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     4321 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     4381 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     4441 atggttcacg tacctagaag ttcctattcc gaagttccta ttctctagaa agtataggaa
     4501 cttccttggc caaaaagcct gaactcaccg cgacgtctgt cgagaagttt ctgatcgaaa
     4561 agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct cgtgctttca
     4621 gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc gatggtttct
     4681 acaaagatcg ttatgtttat cggcactttg catcggccgc gctcccgatt ccggaagtgc
     4741 ttgacattgg ggaattcagc gagagcctga cctattgcat ctcccgccgt gcacagggtg
     4801 tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg gtcgcggagg
     4861 ccatggatgc gatcgctgcg gccgatctta gccagacgag cgggttcggc ccattcggac
     4921 cgcaaggaat cggtcaatac actacatggc gtgattacat atgcgcgatt gctgatcccc
     4981 atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc gcgcaggctc
     5041 tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc gtgcacgcgg
     5101 atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc attgactgga
     5161 gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc tggaggccgt
     5221 ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg gagcttgcag
     5281 gatcgccgcg gctccgggcg tatatgctcc gcattggtct tgaccaactc tatcagagct
     5341 tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac gcaatcgtcc
     5401 gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg gccgtctgga
     5461 ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc actcgtccga
     5521 gggcaaagga atagcacgta ctacgagatt tcgattccac cgccgccttc tatgaaaggt
     5581 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     5641 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     5701 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     5761 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     5821 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5881 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5941 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     6001 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     6061 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6121 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6181 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6241 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6301 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     6361 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     6421 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     6481 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     6541 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     6601 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     6661 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     6721 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6781 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6841 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6901 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6961 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     7021 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     7081 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     7141 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     7201 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     7261 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     7321 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     7381 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     7441 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     7501 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     7561 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     7621 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     7681 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     7741 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7801 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7861 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7921 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     7981 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat
     8041 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc
     8101 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat
     8161 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
     8221 gcgttgacat tgattattga ctaggctttt gcaaaaagct ttgcaaagat ggataaagtt
     8281 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
     8341 t
//
