LOCUS       sequence-345031-        2845 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used to generate IVT RNA for translation extracts.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2845)
  AUTHORS   Mendez AS, Ly M, Gonzalez-Sanchez AM, Hartenian E, Ingolia NT, Cate 
            JH, Glaunsinger BA
  TITLE     The N-terminal domain of SARS-CoV-2 nsp1 plays key roles in 
            suppression of cellular gene expression and preservation of viral 
            gene expression.
  JOURNAL   Cell Rep. 2021 Oct 19;37(3):109841. doi: 
            10.1016/j.celrep.2021.109841. Epub 2021 Sep 30.
  PUBMED    34624207
REFERENCE   2  (bases 1 to 2845)
  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"; journalName: "Cell Rep.";
            date: "2021-10-19"; volume: "37(3):109841. doi"; pages: " 
            10.1016/j.celrep.2021.109841. Epub 2021 Sep 30"
COMMENT     Sequence Label: pUC57-HBB-nLuc
FEATURES             Location/Qualifiers
     source          1..2845
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(188..1003)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(188..1003)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     911..930
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     995..1014
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     1124..1140
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1124..1140
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     terminator      1141..1187
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     1301..1323
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1315..1332
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1316..1332
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1394..1412
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1463..1472
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1469..1981
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             1990..2055
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2121..2137)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2121..2137)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2134..2156)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(join(2309..2845,1..52))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2389..2408)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
       61 tttgatcttt tctacggggt ctgacgctct attcaacaaa gccgccgtcc cgtcaagtca
      121 gcgtaaatgg gtagggggct tcaaatcgtc cgctctgcca gtgttacaac caattaacaa
      181 attctgatta gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat
      241 tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc
      301 agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa
      361 tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag
      421 tgacgactga atccggtgag aatggcaaaa gcttatgcat ttctttccag acttgttcaa
      481 caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc
      541 gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag
      601 gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat
      661 caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc
      721 atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca
      781 gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt
      841 tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt
      901 gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta
      961 atcgcggcct cgagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac
     1021 tgtttatgta agcagacagt tttattgttc atgatgatat atttttatct tgtgcaatgt
     1081 aacatcagag attttgagac acaacgtggc tttcccccgc cgctctagaa ctagtggatc
     1141 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg
     1201 cattatacga gacgtccagg ttgggatacc tgaaacaaaa cccatcgtac ggccaaggaa
     1261 gtctccaata actgtgatcc accacaagcg ccagggtttt cccagtcacg acgttgtaaa
     1321 acgacggcca gtcatgcata atccgcacgc atctggaata aggaagtgcc attccgcctg
     1381 acctccgaat tcgtaatacg actcactata ggacatttgc ttctgacaca actgtgttca
     1441 ctagcaacct caaacagaca ccgccaccat ggtcttcaca ctcgaagatt tcgttgggga
     1501 ctggcgacag acagccggct acaacctgga ccaagtcctt gaacagggag gtgtgtccag
     1561 tttgtttcag aatctcgggg tgtccgtaac tccgatccaa aggattgtcc tgagcggtga
     1621 aaatgggctg aagatcgaca tccatgtcat catcccgtat gaaggtctga gcggcgacca
     1681 aatgggccag atcgaaaaaa tttttaaggt ggtgtaccct gtggatgatc atcactttaa
     1741 ggtgatcctg cactatggca cactggtaat cgacggggtt acgccgaaca tgatcgacta
     1801 tttcggacgg ccgtatgaag gcatcgccgt gttcgacggc aaaaagatca ctgtaacagg
     1861 gaccctgtgg aacggcaaca aaattatcga cgagcgcctg atcaaccccg acggctccct
     1921 gctgttccga gtaaccatca acggagtgac cggctggcgg ctgtgcgaac gcattctggc
     1981 ggcggccgcg attataagga ccacgatggt gactataagg atcacgatat cgattataaa
     2041 gacgacgatg acaagtaggg taccaggcta ggtggaggct cagtgatgat aagtctgcga
     2101 tggtggatgc atgtgtcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaga
     2161 gggcacaatc ctattccgcg ctatccgaca atctccaaga cattaggtgg agttcagttc
     2221 ggcgtatggc atatgtcgct ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     2281 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     2341 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     2401 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     2461 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     2521 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     2581 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     2641 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     2701 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     2761 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     2821 gcaaacaaac caccgctggt agcgg
//
