LOCUS       sequence-403603-        6590 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of full-length SARS-CoV-2 nucleocapsid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6590)
  AUTHORS   Dumelie JG, Chen Q, Miller D, Attarwala N, Gross SS, Jaffrey SR
  TITLE     Biomolecular condensates create phospholipid-enriched 
            microenvironments.
  JOURNAL   Nat Chem Biol. 2023 Nov 16. doi: 10.1038/s41589-023-01474-4.
  PUBMED    37973889
REFERENCE   2  (bases 1 to 6590)
  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: "Nat Chem 
            Biol. 2023 Nov 16. doi: 10.1038/s41589-023-01474-4."
COMMENT     Sequence Label: pET28C-SARS-CoV-2-Nucleocapsid
FEATURES             Location/Qualifiers
     source          1..6590
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             43..60
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(113..131)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      127..174
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      211..666
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(298..317)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     508..529
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(758..1573)
                     /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(758..1573)
                     /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     1481..1500
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1565..1584
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1695..2283
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2184..2203
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2437..2454
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2469..2608
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2534..2553)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2694..2716
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2710..2901)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3710..4792)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     4754..4773
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4793..4870)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5076..5095
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5179..5198
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5179..5197
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5198..5222
                     /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)."
     RBS             5237..5259
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5278..5295
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5305..5322
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             5326..5358
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
ORIGIN
        1 gtccatgagc tcagccgatt ctactcaggc gctctagtcg agcaccacca ccaccaccac
       61 tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
      121 caataactag cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa
      181 ggaggaacta tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
      241 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
      301 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
      361 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
      421 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
      481 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
      541 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
      601 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
      661 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
      721 aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca tcgagcatca
      781 aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
      841 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
      901 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
      961 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     1021 gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat
     1081 cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc
     1141 gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg
     1201 ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg
     1261 ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct
     1321 tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc tcatctgtaa
     1381 catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc
     1441 catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc
     1501 catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac gtttcccgtt
     1561 gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc
     1621 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     1681 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     1741 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     1801 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     1861 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     1921 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     1981 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     2041 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     2101 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     2161 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     2221 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     2281 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     2341 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     2401 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     2461 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat
     2521 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct
     2581 atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc
     2641 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     2701 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag
     2761 ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc
     2821 gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc
     2881 ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt
     2941 aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc
     3001 ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag
     3061 aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg
     3121 tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg
     3181 cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc
     3241 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta
     3301 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg
     3361 cacccgtggg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc
     3421 gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag
     3481 gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc
     3541 cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt
     3601 catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg
     3661 agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc
     3721 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     3781 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca
     3841 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt
     3901 gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt
     3961 cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg
     4021 taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa
     4081 cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc
     4141 cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca
     4201 gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac
     4261 ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac
     4321 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag
     4381 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc
     4441 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca
     4501 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt
     4561 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc
     4621 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca
     4681 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct
     4741 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca
     4801 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt
     4861 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc
     4921 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg
     4981 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
     5041 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
     5101 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
     5161 atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
     5221 cccctctaga aataattttg tttaacttta agaaggagat ataccatggg cagcagccat
     5281 catcatcatc atcacagcag cggcctggtg ccgcgcggca gccatatggc tagcatgact
     5341 ggtggacagc aaatgggtcg gatcatgagc gataacggcc cccaaaacca acggaatgcc
     5401 cctcggataa catttggtgg ccctagtgat tccacaggaa gtaaccaaaa cggagaacgg
     5461 tctggagccc gctctaaaca gaggcgacca cagggattgc caaataacac agcgtcatgg
     5521 tttacagccc tgactcaaca tggaaaggag gacttgaaat ttccacgagg acagggtgta
     5581 ccaataaaca caaacagtag tccagacgat cagatcgggt attatagaag ggccacgaga
     5641 cgcatcagag gtggagatgg taaaatgaaa gatctcagtc cgcgctggta cttttattat
     5701 ttgggaaccg gcccagaagc gggcctccca tatggagcca ataaggacgg gattatatgg
     5761 gtggctacag agggagccct caatacgcca aaagatcaca tcggaacaag aaaccctgct
     5821 aataatgccg caatcgtgct gcagttgcct caggggacga cgcttcctaa aggcttttac
     5881 gcagaaggat cacgcggcgg cagccaagca tcctcaaggt ctagttccag aagtcgaaac
     5941 agctccagaa actccacacc agggtccagt aggggcacaa gtccggcgcg gatggcgggc
     6001 aacggcggag acgccgcact cgctctgttg ctcctggacc ggctcaacca acttgaatcc
     6061 aagatgagtg gtaagggaca gcagcaacaa ggtcaaaccg taaccaagaa aagcgctgca
     6121 gaagcttcta aaaaacctcg acaaaaacgg accgctacga aggcatataa cgttactcaa
     6181 gccttcggga gacgagggcc ggagcaaacc caggggaatt ttggagatca ggagctcatc
     6241 cggcaaggga cagattacaa acattggcct caaatcgctc agttcgcgcc gtccgcaagt
     6301 gcttttttcg gaatgtctcg catcggtatg gaagtaacac catccgggac atggctcact
     6361 tataccggcg ccatcaagct ggacgacaaa gatccaaatt tcaaagatca ggtgattctc
     6421 ttgaataaac acatagacgc gtataagacg tttccaccca ccgaacccaa gaaggacaaa
     6481 aagaaaaaag ctgatgaaac acaagcgttg ccccagcgac agaagaagca gcaaaccgtc
     6541 accctcctgc cagccgctga tctcgatgac ttctccaagc aacttcaaca
//
