LOCUS       sequence-476565-        8183 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes full-length human TMEM106B. M210A/F213A mutations block S1 
            binding and SARS-CoV-2 Belgium/GHB-03021/2020 TMEM106B mediated 
            infection, without affecting TMEM106B expression or localization..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8183)
  AUTHORS   Baggen J, Jacquemyn M, Persoons L, Vanstreels E, Pye VE, Wrobel AG, 
            Calvaresi V, Martin SR, Roustan C, Cronin NB, Reading E, Thibaut HJ,
            Vercruysse T, Maes P, De Smet F, Yee A, Nivitchanyong T, Roell M, 
            Franco-Hernandez N, Rhinn H, Mamchak AA, Ah Young-Chapon M, Brown E,
            Cherepanov P, Daelemans D
  TITLE     TMEM106B is a receptor mediating ACE2-independent SARS-CoV-2 cell 
            entry.
  JOURNAL   Cell. 2023 Aug 3;186(16):3427-3442.e22. doi: 
            10.1016/j.cell.2023.06.005. Epub 2023 Jul 7.
  PUBMED    37421949
REFERENCE   2  (bases 1 to 8183)
  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: 
            "10.1016/j.cell.2023.06.005"; journalName: "Cell"; date: "2023-08-3-
            3"; volume: "186"; issue: "16"; pages: "3427-3442.e22"
COMMENT     Sequence Label: pLCKO-CMV-TMEM106B-M210A_F213A-P2A-Blasticidin
FEATURES             Location/Qualifiers
     source          1..8183
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     718..742
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      840..849
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1677..1733
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1734..2129
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    2145..2733
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2198..2218)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2616..2627)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2805..3038
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3116..3237
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3153..3172)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3207..3226
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3277..3412
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3432..3451)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3433..3451)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3461..3478)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3461..3477)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3470..3492)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3619..4074
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3706..3725)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3916..3937
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4100..4204
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4205..4273,4274..5065)
                     /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     complement(4423..4442)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5236..5824
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5725..5744
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5978..5995
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6112..6133
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6148..6153,6154..6171,6172..6178)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6186..6202
                     /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)."
     primer_bind     6191..6213
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6210..6226
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6210..6226
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6245..6265
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6247..6265
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6293..6519
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6520..6700
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6747..6872
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7365..7598
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7783..7827
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    8014..8131
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tcgtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgggc ctcggcggcc aagcttggca atccggtact gttggtaaag
      841 ccaccatggg aaagtctctt tctcatttgc ctttgcattc aagcaaagaa gatgcttatg
      901 atggtgtaac gtcagagaat atgcgcaatg gactggttaa tagtgaagtc cataatgaag
      961 atggaagaaa tggagatgtc tctcagtttc catatgtgga atttacagga agagatagtg
     1021 tcacctgccc tacttgtcag ggtactggtc gtatcccacg cgggcaagaa aaccaactgg
     1081 tggcattgat tccatatagt gatcagagat taaggccaag aagaacaaag ctgtatgtga
     1141 tggcttctgt gtttgtctgt ctactccttt ctggattggc tgtgtttttc cttttccctc
     1201 ggtcaattga tgtcaagtac attggtgtaa aatcagccta tgtcagttat gatgttcaga
     1261 agcgtacaat ttatttaaat atcacaaaca cactaaatat aacaaacaat aactattact
     1321 ctgtcgaagt tgaaaacatc actgcccaag ttcaattttc gaagactgta atcggcaaag
     1381 cacgcttaaa caacataacc attattggtc cacttgatat gaaacaaatt gattacacag
     1441 tacctaccgt tatagcagag gaaatgagtt atgcgtatga tgcctgtact ctgatatcca
     1501 tcaaagtgca taacatagta ctcatgatgc aagttactgt gacaacaaca tactttggcc
     1561 actctgaaca gatatcccag gagaggtatc agtatgtcga ctgtggaaga aacacaactt
     1621 atcagttggg gcagtctgaa tatttaaatg tacttcagcc acaacagggc agcggagcaa
     1681 ccaattttag tctcctcaaa caggccggcg atgtcgaaga aaatccgggc ccagccaagc
     1741 ctttgtctca agaagaatcc accctcattg aaagagcaac ggctacaatc aacagcatcc
     1801 ccatctctga agactacagc gtcgccagcg cagctctctc tagcgacggc cgcatcttca
     1861 ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc agaactcgtg gtgctgggca
     1921 ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt cgcgatcgga aatgagaaca
     1981 ggggcatctt gagcccctgc ggacggtgcc gacaggtgct tctcgatctg catcctggga
     2041 tcaaagccat agtgaaggac agtgatggac agccgacggc agttgggatt cgtgaattgc
     2101 tgccctctgg ttatgtgtgg gagggctgaa cgcgttaagt cgacaatcaa cctctggatt
     2161 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     2221 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     2281 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     2341 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     2401 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     2461 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     2521 ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt gttgccacct
     2581 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     2641 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     2701 cgagtcggat ctccctttgg gccgcctccc cgcgtcgact ttaagaccaa tgacttacaa
     2761 ggcagctgta gatcttagcc actttttaaa agaaaagggg ggactggaag ggctaattca
     2821 ctcccaacga agacaagatc tgctttttgc ttgtactggg tctctctggt tagaccagat
     2881 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
     2941 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
     3001 cctcagaccc ttttagtcag tgtggaaaat ctctagcagt acgtatagta gttcatgtca
     3061 tcttattatt cagtatttat aacttgcaaa gaaatgaata tcagagagtg agaggaactt
     3121 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     3181 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     3241 tgtctggctc tagctatccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     3301 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     3361 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctagggacg
     3421 tacccaattc gccctatagt gagtcgtatt acgcgcgctc actggccgtc gttttacaac
     3481 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
     3541 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
     3601 gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     3661 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     3721 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     3781 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     3841 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     3901 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     3961 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     4021 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
     4081 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     4141 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     4201 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     4261 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     4321 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     4381 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     4441 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     4501 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     4561 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     4621 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     4681 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     4741 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     4801 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     4861 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     4921 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     4981 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     5041 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     5101 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     5161 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     5221 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     5281 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     5341 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     5401 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     5461 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     5521 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     5581 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     5641 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     5701 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     5761 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     5821 gaaaaacgcc agcaacgcgg cctttttacg gttcctggtc ttttgctggc cttttgctca
     5881 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     5941 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     6001 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     6061 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
     6121 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
     6181 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
     6241 gcgcgcaatt aaccctcact aaagggaaca aaagctggag ctgcaagctt aatgtagtct
     6301 tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca tgccttacaa
     6361 ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa ggtggtacga tcgtgcctta
     6421 ttaggaaggc aacagacggg tctgacatgg attggacgaa ccactgaatt gccgcattgc
     6481 agagatattg tatttaagtg cctagctcga tacataaacg ggtctctctg gttagaccag
     6541 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
     6601 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
     6661 tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg aacagggact
     6721 tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt gctgaagcgc
     6781 gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg actagcggag
     6841 gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga attagatcgc
     6901 gatgggaaaa aattcggtta aggccagggg gaaagaaaaa atataaatta aaacatatag
     6961 tatgggcaag cagggagcta gaacgattcg cagttaatcc tggcctgtta gaaacatcag
     7021 aaggctgtag acaaatactg ggacagctac aaccatccct tcagacagga tcagaagaac
     7081 ttagatcatt atataataca gtagcaaccc tctattgtgt gcatcaaagg atagagataa
     7141 aagacaccaa ggaagcttta gacaagatag aggaagagca aaacaaaagt aagaccaccg
     7201 cacagcaagc ggccgctgat cttcagacct ggaggaggag atatgaggga caattggaga
     7261 agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc acccaccaag
     7321 gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc tttgttcctt
     7381 gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct gacggtacag
     7441 gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag ggctattgag
     7501 gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca ggcaagaatc
     7561 ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg ttgctctgga
     7621 aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa atctctggaa
     7681 cagatttgga atcacacgac ctggatggag tgggacagag aaattaacaa ttacacaagc
     7741 ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga acaagaatta
     7801 ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa ttggctgtgg
     7861 tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat agtttttgct
     7921 gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt tcagacccac
     7981 ctcccaaccc cgaggggacc cagagagggc ctattttaaa agaaaagggg ggattggggg
     8041 gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa ctaaagaatt
     8101 acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca gcagagatcc
     8161 actttggcgc cggctcgagg ggg
//
