LOCUS       sequence-376741-        7090 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 7090)
  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 7090)
  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: CPSF4
FEATURES             Location/Qualifiers
     source          1..7090
                     /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(2047..2071)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2196..2237
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2692..2709)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2698..2922
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3055..3074)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3206..3253
                     /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             3261..4283
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3261..4283
                     /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    4413..4534
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4450..4469)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4504..4523
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4583..4599)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4583..4599)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4596..4618)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4607..4623
                     /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(4631..4637,4638..4655,4656..4661))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4676..4697
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4814..4831)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4985..5573)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5065..5084)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5744..6535,6536..6604))
                     /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     6367..6386
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6605..6709)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6784..6803)
                     /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 gttggcatgc
     1321 aggaaatcat cgccagcgtg gaccacatca agtttgactt ggagatcgcg gtggagcagc
     1381 agctgggggc gcagccgctg cccttccccg gcatggacaa gtcgggcgct gctgtctgtg
     1441 aattcttttt gaaagctgcc tgcggcaaag ggggcatgtg tccgtttcgc cacatcagtg
     1501 gtgagaagac agttgtgtgc aaacactggc tgcgtggcct atgcaagaaa ggggaccagt
     1561 gtgagttcct gcatgagtat gacatgacca agatgcccga gtgctacttc tactccaagt
     1621 tcggggagtg cagcaacaag gaatgtccct tcctgcacat cgaccccgag tccaagatca
     1681 aggactgtcc ttggtatgac cgcggcttct gcaagcacgg tcccctctgc aggcaccggc
     1741 acacacggag agtcatctgt gtgaattacc tcgtgggatt ctgcccggag gggccctcgt
     1801 gtaaattcat gcaccctcga tttgaactgc ccatgggaac caccgagcag cccccactgc
     1861 cacagcagac acagcctcca gcaaagagaa ccccgcaggt catcggggtc atgcagagtc
     1921 aaaacagcag cgcgggcaac cggggacccc ggccactgga gcaggtcacc tgttacaagt
     1981 gtggcgagaa aggacactac gccaacagat gcaccaaagg gcacttggcc tttctcagtg
     2041 gacagtgccc aactttcttg tacaaagtgg ttgatatcca gcacagtggc ggccgctcga
     2101 gtctagaggg cccgcggttc gaaaccagcg gaggcggcgg aagcggggga ggaggcagcg
     2161 gcggaggcgg cagcgggggc ggcggaagcg ctaccggtaa gcctatccct aaccctctcc
     2221 tcggtctcga ttctacgcgt accggtatgg cttctaactt tactcagttc gttctcgtcg
     2281 acaatggcgg aactggcgac gtgactgtcg ccccaagcaa cttcgctaac ggggtcgctg
     2341 aatggatcag ctctaactcg cgatcacagg cttacaaagt aacctgtagc gttcgtcaga
     2401 gctctgcgca gaatcgcaaa tacaccatca aagtcgaggt gcctaaagtg gcaacccaga
     2461 ctgttggtgg tgaagagctt cctgtagccg gatggagatc ttacttaaat atggaactaa
     2521 ccattccaat tttcgccacg aattccgact gcgagcttat tgttaaggca atgcaaggtc
     2581 tcctaaaaga tggaaacccg attccctcgg ccatcgcggc caactccggc atctacggag
     2641 gtggaggtgg atctcgtacg ggttagtaat gagtttaaac ccgctgatca gcctcgactg
     2701 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     2761 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     2821 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     2881 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     2941 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     3001 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     3061 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     3121 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     3181 attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat tctctagaaa
     3241 gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc
     3301 tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc gaagaatctc
     3361 gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg
     3421 atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg ctcccgattc
     3481 cggaagtgct tgacattggg gaattcagcg agagcctgac ctattgcatc tcccgccgtg
     3541 cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg
     3601 tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc gggttcggcc
     3661 cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata tgcgcgattg
     3721 ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt gcgtccgtcg
     3781 cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc cggcacctcg
     3841 tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata acagcggtca
     3901 ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac atcttcttct
     3961 ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg aggcatccgg
     4021 agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt gaccaactct
     4081 atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg
     4141 caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg
     4201 ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga cgccccagca
     4261 ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc gccgccttct
     4321 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     4381 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     4441 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4501 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4561 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4621 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4681 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4741 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4801 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4861 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4921 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4981 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5041 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5101 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5161 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5221 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5281 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5341 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5401 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5461 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5521 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5581 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5641 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5701 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5761 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5821 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5881 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5941 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     6001 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     6061 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     6121 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     6181 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6241 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6301 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6361 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6421 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6481 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6541 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6601 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6661 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6721 gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct
     6781 cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt
     6841 ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga
     6901 caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc
     6961 cagatatacg cgttgacatt gattattgac taggcttttg caaaaagctt tgcaaagatg
     7021 gataaagttt taaacagaga ggaatctttg cagctaatgg accttctagg tcttgaaagg
     7081 agtgggaatt
//
