LOCUS       sequence-317328-        7939 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 7939)
  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 7939)
  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: CCNL1
FEATURES             Location/Qualifiers
     source          1..7939
                     /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"
     regulatory      2855..2864
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(2896..2920)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3045..3086
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3541..3558)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3547..3771
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3904..3923)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4055..4102
                     /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             4110..5132
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4110..5132
                     /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    5262..5383
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5299..5318)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5353..5372
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5432..5448)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5432..5448)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5445..5467)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5456..5472
                     /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(5480..5486,5487..5504,5505..5510))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5525..5546
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5663..5680)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5834..6422)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5914..5933)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6593..7384,7385..7453))
                     /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     7216..7235
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7454..7558)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7633..7652)
                     /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 gttggcatgg
     1321 cgtccgggcc tcattcgaca gctactgctg ccgcagccgc ctcatcggcc gccccaagcg
     1381 cgggcggctc cagctccggg acgacgacca cgacgacgac cacgacggga gggatcctga
     1441 tcggcgatcg cctgtactcg gaagtttcac ttaccatcga ccactctctg attccggagg
     1501 agaggctctc gcccacccca tccatgcagg atgggctcga cctgcccagt gagacggact
     1561 tacgcatcct gggctgcgag ctcatccagg ccgccggcat tctcctccgg ctgccgcagg
     1621 tggcgatggc aacggggcag gtgttgtttc atcgtttttt ctactccaaa tctttcgtca
     1681 aacacagttt cgagattgtt gctatggctt gtattaatct tgcatcaaaa atcgaagaag
     1741 cacctagaag aataagagat gtgattaatg tattccacca cctccgccag ttaagaggaa
     1801 aaaggactcc aagccccctg atccttgatc agaactacat taacaccaaa aatcaagtta
     1861 tcaaagcaga gaggagggtg ctaaaggagt tgggattttg tgttcatgtc aagcatcctc
     1921 ataagatcat tgttatgtat ttacaagtct tagaatgtga acgtaatcaa accctggttc
     1981 aaactgcctg gaattacatg aatgacagtc ttcgaaccaa tgtgtttgtt cgatttcaac
     2041 cagagactat agcatgtgct tgcatctacc ttgcagctag agcacttcag attccgttgc
     2101 caactcgtcc ccattggttt cttctttttg gtactacaga agaggaaatc caggaaatct
     2161 gcatagaaac acttaggctt tataccagaa aaaagccaaa ctatgaatta ctggaaaaag
     2221 aagtagaaaa aagaaaagta gccttacaag aagccaaatt aaaagcaaag ggattgaatc
     2281 cggatggaac tccagccctt tcaaccctgg gtggattttc tccagcctcc aagccatcat
     2341 caccaagaga agtaaaagct gaagagaaat caccaatctc cattaatgtg aagacagtca
     2401 aaaaagaacc tgaggataga caacaggctt ccaaaagccc ttacaatggt gtaagaaaag
     2461 acagcaagag aagtagaaat agcagaagtg caagtcgatc gaggtcaaga acacgatcac
     2521 gttctagatc acatactcca agaagacact ataataatag gcggagtcga tctggaacat
     2581 acagctcgag atcaagaagc aggtcccgca gtcacagtga aagccctcga agacatcata
     2641 atcatggttc tcctcacctt aaggccaagc ataccagaga tgatttaaaa agttcaaaca
     2701 gacatggtca taaaaggaaa aaatctcgtt ctcgatctca gagcaagtct cgggatcact
     2761 cagatgcagc caagaaacac aggcatgaaa ggggacatca tagggacagg cgtgaacgat
     2821 ctcgctcctt tgagaggtcc cataaaagca agcaccatgg tggcagtcgc tcaggacatg
     2881 gcaggcacag gcgctgccca actttcttgt acaaagtggt tgatatccag cacagtggcg
     2941 gccgctcgag tctagagggc ccgcggttcg aaaccagcgg aggcggcgga agcgggggag
     3001 gaggcagcgg cggaggcggc agcgggggcg gcggaagcgc taccggtaag cctatcccta
     3061 accctctcct cggtctcgat tctacgcgta ccggtatggc ttctaacttt actcagttcg
     3121 ttctcgtcga caatggcgga actggcgacg tgactgtcgc cccaagcaac ttcgctaacg
     3181 gggtcgctga atggatcagc tctaactcgc gatcacaggc ttacaaagta acctgtagcg
     3241 ttcgtcagag ctctgcgcag aatcgcaaat acaccatcaa agtcgaggtg cctaaagtgg
     3301 caacccagac tgttggtggt gaagagcttc ctgtagccgg atggagatct tacttaaata
     3361 tggaactaac cattccaatt ttcgccacga attccgactg cgagcttatt gttaaggcaa
     3421 tgcaaggtct cctaaaagat ggaaacccga ttccctcggc catcgcggcc aactccggca
     3481 tctacggagg tggaggtgga tctcgtacgg gttagtaatg agtttaaacc cgctgatcag
     3541 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     3601 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     3661 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     3721 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     3781 cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa
     3841 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     3901 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     3961 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4021 aaaaacttga ttagggtgat ggttcacgta cctagaagtt cctattccga agttcctatt
     4081 ctctagaaag tataggaact tccttggcca aaaagcctga actcaccgcg acgtctgtcg
     4141 agaagtttct gatcgaaaag ttcgacagcg tctccgacct gatgcagctc tcggagggcg
     4201 aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata
     4261 gctgcgccga tggtttctac aaagatcgtt atgtttatcg gcactttgca tcggccgcgc
     4321 tcccgattcc ggaagtgctt gacattgggg aattcagcga gagcctgacc tattgcatct
     4381 cccgccgtgc acagggtgtc acgttgcaag acctgcctga aaccgaactg cccgctgttc
     4441 tgcagccggt cgcggaggcc atggatgcga tcgctgcggc cgatcttagc cagacgagcg
     4501 ggttcggccc attcggaccg caaggaatcg gtcaatacac tacatggcgt gatttcatat
     4561 gcgcgattgc tgatccccat gtgtatcact ggcaaactgt gatggacgac accgtcagtg
     4621 cgtccgtcgc gcaggctctc gatgagctga tgctttgggc cgaggactgc cccgaagtcc
     4681 ggcacctcgt gcacgcggat ttcggctcca acaatgtcct gacggacaat ggccgcataa
     4741 cagcggtcat tgactggagc gaggcgatgt tcggggattc ccaatacgag gtcgccaaca
     4801 tcttcttctg gaggccgtgg ttggcttgta tggagcagca gacgcgctac ttcgagcgga
     4861 ggcatccgga gcttgcagga tcgccgcggc tccgggcgta tatgctccgc attggtcttg
     4921 accaactcta tcagagcttg gttgacggca atttcgatga tgcagcttgg gcgcagggtc
     4981 gatgcgacgc aatcgtccga tccggagccg ggactgtcgg gcgtacacaa atcgcccgca
     5041 gaagcgcggc cgtctggacc gatggctgtg tagaagtact cgccgatagt ggaaaccgac
     5101 gccccagcac tcgtccgagg gcaaaggaat agcacgtact acgagatttc gattccaccg
     5161 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     5221 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     5281 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     5341 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     5401 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     5461 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     5521 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     5581 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     5641 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     5701 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5761 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5821 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     5881 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     5941 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     6001 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     6061 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6121 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     6181 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6241 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     6301 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     6361 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     6421 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     6481 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     6541 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     6601 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     6661 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     6721 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     6781 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     6841 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     6901 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     6961 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     7021 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     7081 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     7141 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     7201 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     7261 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     7321 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     7381 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     7441 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     7501 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     7561 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
     7621 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
     7681 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
     7741 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
     7801 tgtacgggcc agatatacgc gttgacattg attattgact aggcttttgc aaaaagcttt
     7861 gcaaagatgg ataaagtttt aaacagagag gaatctttgc agctaatgga ccttctaggt
     7921 cttgaaagga gtgggaatt
//
