LOCUS       sequence-376939-        7888 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 7888)
  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 7888)
  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: NAP1L3
FEATURES             Location/Qualifiers
     source          1..7888
                     /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(2845..2869)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2994..3035
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3490..3507)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3496..3720
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3853..3872)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4004..4051
                     /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             4059..5081
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4059..5081
                     /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    5211..5332
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5248..5267)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5302..5321
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5381..5397)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5381..5397)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5394..5416)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5405..5421
                     /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(5429..5435,5436..5453,5454..5459))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5474..5495
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5612..5629)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5783..6371)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5863..5882)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6542..7333,7334..7402))
                     /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     7165..7184
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7403..7507)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7582..7601)
                     /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 ccatggcaga agcagatttt aaaatggtct cggaacctgt cgcccatggg gttgccgaag
     1381 aggagatggc tagctcgact agtgattctg gggaagaatc tgacagcagt agctctagca
     1441 gcagcactag tgacagcagc agcagcagca gcactagtgg cagcagcagc ggcagcggca
     1501 gcagcagcag cagcagcggc agcactagca gccgcagccg cttgtataga aagaagaggg
     1561 tacctgagcc ttccagaagg gcgcggcggg ccccgttggg aacaaatttc gtggataggc
     1621 tgcctcaggc agttagaaat cgtgtgcaag cgcttagaaa cattcaagat gaatgtgaca
     1681 aggtagatac cctgttctta aaagcaattc atgatcttga aagaaaatat gctgaactca
     1741 acaagcctct gtatgatagg cggtttcaaa tcatcaatgc agaatacgag cctacagaag
     1801 aagaatgtga atggaattca gaggatgagg agttcagcag tgatgaggag gtgcaggata
     1861 acacccctag tgaaatgcct cccttagagg gtgaggaaga agaaaaccct aaagaaaacc
     1921 cagaggtgaa agctgaagag aaggaagttc ctaaagaaat tcctgaggtg aaggatgaag
     1981 aaaaggaagt tgctaaagaa attactgagg taaaggctga agaaaaagca gattctaaag
     2041 actgtatgga ggcaacccct gaagtaaaag aagatcctaa agaagtcccc caggtaaagg
     2101 cagatgataa agaacagcct aaagcaacag aggctaaggc aagggctgca gtaagagaga
     2161 ctcataaaag agttcctgag gaaaggcttc aggacagtgt agatcttaaa agagctagga
     2221 agggaaagcc taaaagagaa gaccctaaag gcattcctga ctattggctg attgttttaa
     2281 agaatgttga caagctcggg cctatgattc agaagtatga tgagcccatt ctgaagttct
     2341 tgtcggatgt tagcctgaag ttctcaaaac ctggccagcc tgtaagttac acctttgaat
     2401 ttcattttct acccaaccca tacttcagaa atgaggtgct ggtgaagaca tatataataa
     2461 aggcaaaacc agatcacaat gatcccttct tttcttgggg atgggaaatt gaagattgca
     2521 aaggctgcaa gatagactgg agaagaggaa aagatgttac tgtgacaact acccagagtc
     2581 gcacaactgc tactggagaa attgaaatcc agccaagagt ggttcctaat gcatcattct
     2641 tcaacttctt tagtcctcct gagattccta tgattgggaa gctggaacca cgagaagatg
     2701 ctatcctgga tgaggacttt gaaattgggc agattttaca tgataatgtc atcctgaaat
     2761 caatctatta ctatactgga gaagtcaatg gtacctacta tcaatttggc aaacattatg
     2821 gaaacaagaa atacagaaaa ttggacccag ctttcttgta caaagtggtt gatatccagc
     2881 acagtggcgg ccgctcgagt ctagagggcc cgcggttcga aaccagcgga ggcggcggaa
     2941 gcgggggagg aggcagcggc ggaggcggca gcgggggcgg cggaagcgct accggtaagc
     3001 ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac cggtatggct tctaacttta
     3061 ctcagttcgt tctcgtcgac aatggcggaa ctggcgacgt gactgtcgcc ccaagcaact
     3121 tcgctaacgg ggtcgctgaa tggatcagct ctaactcgcg atcacaggct tacaaagtaa
     3181 cctgtagcgt tcgtcagagc tctgcgcaga atcgcaaata caccatcaaa gtcgaggtgc
     3241 ctaaagtggc aacccagact gttggtggtg aagagcttcc tgtagccgga tggagatctt
     3301 acttaaatat ggaactaacc attccaattt tcgccacgaa ttccgactgc gagcttattg
     3361 ttaaggcaat gcaaggtctc ctaaaagatg gaaacccgat tccctcggcc atcgcggcca
     3421 actccggcat ctacggaggt ggaggtggat ctcgtacggg ttagtaatga gtttaaaccc
     3481 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     3541 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     3601 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     3661 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     3721 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
     3781 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     3841 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     3901 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     3961 tcgaccccaa aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa
     4021 gttcctattc tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga
     4081 cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct
     4141 cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc
     4201 gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat
     4261 cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct
     4321 attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc
     4381 ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc
     4441 agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg
     4501 atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca
     4561 ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc
     4621 ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg
     4681 gccgcataac agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg
     4741 tcgccaacat cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact
     4801 tcgagcggag gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca
     4861 ttggtcttga ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg
     4921 cgcagggtcg atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa
     4981 tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg
     5041 gaaaccgacg ccccagcact cgtccgaggg caaaggaata gcacgtacta cgagatttcg
     5101 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     5161 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     5221 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     5281 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     5341 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     5401 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     5461 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     5521 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     5581 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     5641 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     5701 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     5761 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     5821 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     5881 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     5941 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6001 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6061 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6121 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     6181 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     6241 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     6301 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     6361 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     6421 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     6481 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     6541 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6601 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6661 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6721 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     6781 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6841 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6901 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6961 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     7021 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     7081 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     7141 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     7201 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     7261 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     7321 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     7381 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     7441 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     7501 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga
     7561 tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     7621 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
     7681 aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct
     7741 gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta ggcttttgca
     7801 aaaagctttg caaagatgga taaagtttta aacagagagg aatctttgca gctaatggac
     7861 cttctaggtc ttgaaaggag tgggaatt
//
