LOCUS       sequence-345027-        6543 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses delta 122-130 CoV2 nsp1 in mammalian cells. This contains 
            a 3XFLAG Halo tag fusion to nsp1..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6543)
  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 6543)
  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: pCDNA4 3xFLAG Halo delta 122-130 nsp1
FEATURES             Location/Qualifiers
     source          1..6543
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    586..604
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    607..625
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     635..659
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             760..825
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             835..1722
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="AEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     CDS             1735..1755
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     primer_bind     complement(2317..2334)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2323..2547
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2593..3021
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2680..2699)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2890..2911
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3030..3050)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3035..3364
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3215..3350
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3277..3296
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3412..3459
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3478..3852
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    3982..4103
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4019..4038)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4073..4092
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4152..4168)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4152..4168)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4165..4187)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4176..4192
                     /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(4200..4206,4207..4224,4225..4230))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4245..4266
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4383..4400)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4554..5142)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4634..4653)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5313..6104,6105..6173))
                     /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     5936..5955
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6174..6278)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6353..6372)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataagga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctccct atcagtgata
      601 gagatctccc tatcagtgat agagatcgtc gacgagctcg tttagtgaac cgtcagatcg
      661 cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc
      721 tccggactct agcgtttaaa cttaagctta ccaatgatgg actacaagga ccacgacggc
      781 gactacaagg accacgacat cgactacaag gacgacgacg acaagggggc ggccgcagaa
      841 atcggtactg gctttccatt cgacccccat tatgtggaag tcctgggcga gcgcatgcac
      901 tacgtcgatg ttggtccgcg cgatggcacc cctgtgctgt tcctgcacgg taacccgacc
      961 tcctcctacg tgtggcgcaa catcatcccg catgttgcac cgacccatcg ctgcattgct
     1021 ccagacctga tcggtatggg caaatccgac aaaccagacc tgggttattt cttcgacgac
     1081 cacgtccgct tcatggatgc cttcatcgaa gccctgggtc tggaagaggt cgtcctggtc
     1141 attcacgact ggggctccgc tctgggtttc cactgggcca agcgcaatcc agagcgcgtc
     1201 aaaggtattg catttatgga gttcatccgc cctatcccga cctgggacga atggccagaa
     1261 tttgcccgcg agaccttcca ggccttccgc accaccgacg tcggccgcaa gctgatcatc
     1321 gatcagaacg tttttatcga gggtacgctg ccgatgggtg tcgtccgccc gctgactgaa
     1381 gtcgagatgg accattaccg cgagccgttc ctgaatcctg ttgaccgcga gccactgtgg
     1441 cgcttcccaa acgagctgcc aatcgccggt gagccagcga acatcgtcgc gctggtcgaa
     1501 gaatacatgg actggctgca ccagtcccct gtcccgaagc tgctgttctg gggcacccca
     1561 ggcgttctga tcccaccggc cgaagccgct cgcctggcca aaagcctgcc taactgcaag
     1621 gctgtggaca tcggcccggg tctgaatctg ctgcaagaag acaacccgga cctgatcggc
     1681 agcgagatcg cgcgctggct gtcgacgctc gagatttccg gcgagccaac cactgaggat
     1741 ctgtactttc agagcgcggc cgagagcctt gtccctggtt tcaacgagaa aacacacgtc
     1801 caactcagtt tgcctgtttt acaggttcgc gacgtgctcg tacgtggctt tggagactcc
     1861 gtggaggagg tcttatcaga ggcacgtcaa catcttaaag atggcacttg tggcttagta
     1921 gaagttgaaa aaggcgtttt gcctcaactt gaacagccct atgtgttcat caaacgttcg
     1981 gatgctcgaa ctgcacctca tggtcatgtt atggttgagc tggtagcaga actcgaaggc
     2041 attcagtacg gtcgtagtgg tgagacactt ggtgtccttg tccctcatgt gggcgaaata
     2101 ccagtggctt accgcaaggt tgctggtggc catagttacg gcgccgatct aaagtcattt
     2161 gacttaggcg acgagcttgg cactgatcct tatgaagatt ttcaagaaaa ctggaacact
     2221 aaacatagca gtggtgttac ccgtgaactc atgcgtgagc ttaacggagg gtagggccgc
     2281 tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc
     2341 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     2401 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     2461 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     2521 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     2581 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     2641 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     2701 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     2761 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     2821 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     2881 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     2941 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     3001 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     3061 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     3121 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     3181 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     3241 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     3301 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     3361 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat
     3421 taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg
     3481 gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag
     3541 ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg
     3601 gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac
     3661 accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc
     3721 gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg
     3781 tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag
     3841 gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     3901 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     3961 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     4021 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     4081 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     4141 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     4201 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     4261 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4321 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     4381 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     4441 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4501 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4561 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4621 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4681 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4741 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4801 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4861 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4921 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4981 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     5041 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     5101 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     5161 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     5221 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     5281 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     5341 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     5401 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     5461 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     5521 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5581 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5641 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5701 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5761 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5821 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5881 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5941 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     6001 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     6061 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     6121 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     6181 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     6241 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     6301 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
     6361 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
     6421 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
     6481 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
     6541 gtt
//
