LOCUS       sequence-345024-        6381 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses delta 118-180 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 6381)
  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 6381)
  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 118-180 nsp1
FEATURES             Location/Qualifiers
     source          1..6381
                     /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(2155..2172)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2161..2385
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2431..2859
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2518..2537)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2728..2749
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2868..2888)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2873..3202
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3053..3188
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3115..3134
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3250..3297
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3316..3690
                     /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    3820..3941
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3857..3876)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3911..3930
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3990..4006)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3990..4006)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4003..4025)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4014..4030
                     /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(4038..4044,4045..4062,4063..4068))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4083..4104
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4221..4238)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4392..4980)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4472..4491)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5151..5942,5943..6011))
                     /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     5774..5793
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6012..6116)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6191..6210)
                     /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 ccaataggga
      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 agggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga
     2161 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     2221 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2281 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2341 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     2401 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     2461 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2521 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2581 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2641 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2701 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2761 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2821 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     2881 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2941 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3001 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3061 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3121 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     3181 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     3241 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     3301 gtgaggaact aaaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac
     3361 gtcgccggag cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag
     3421 gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac
     3481 caggtggtgc cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac
     3541 gccgagtggt cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc
     3601 gagatcggcg agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc
     3661 gtgcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga ttccaccgcc
     3721 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     3781 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     3841 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3901 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     3961 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4021 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4081 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4141 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4201 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4261 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4321 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4381 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4441 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4501 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4561 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4621 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4681 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4741 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4801 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4861 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4921 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4981 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     5041 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     5101 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     5161 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     5221 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     5281 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     5341 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     5401 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     5461 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     5521 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     5581 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5641 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5701 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5761 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5821 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5881 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5941 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     6001 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     6061 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     6121 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
     6181 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
     6241 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
     6301 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
     6361 tacgggccag atatacgcgt t
//
