LOCUS       sequence-417565-        6662 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TRAM2 fused with 2Strep in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6662)
  AUTHORS   Li J, Gui Q, Liang FX, Sall J, Zhang Q, Duan Y, Dhabaria A, Askenazi
            M, Ueberheide B, Stapleford KA, Pagano M
  TITLE     The REEP5/TRAM1 complex binds SARS-CoV-2 NSP3 and promotes virus 
            replication.
  JOURNAL   J Virol. 2023 Sep 28:e0050723. doi: 10.1128/jvi.00507-23.
  PUBMED    37768083
REFERENCE   2  (bases 1 to 6662)
  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: "J Virol. 
            2023 Sep 28:e0050723. doi: 10.1128/jvi.00507-23."
COMMENT     Sequence Label: TRAM2-2Strep
FEATURES             Location/Qualifiers
     source          1..6662
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(35..54)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        226..605
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        606..809
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     760..780
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     854..873
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        854..872
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2023..2032
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2027..2036
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2044..2067
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             2104..2127
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             2152..2175
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     promoter        complement(2206..2224)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2207..2224)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2244..2261)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2250..2474
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2520..2948
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2607..2626)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2817..2838
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2957..2977)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2962..3291
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3142..3277
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3204..3223
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3358..4152
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3412..3431)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4022..4041
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4326..4447
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4363..4382)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4417..4436
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4496..4512)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4496..4512)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4509..4531)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4520..4536
                     /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(4544..4550,4551..4568,4569..4574))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4589..4610
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4727..4744)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4898..5486)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4978..4997)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5657..6448,6449..6517))
                     /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     6280..6299
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6518..6622)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
       61 taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa
      121 tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg cttagggtta
      181 ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat tgattattga
      241 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
      301 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
      361 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      421 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      481 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      541 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      601 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
      661 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
      721 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
      781 tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc actgcttact
      841 ggcttatcga aattaatacg actcactata gggagaccca agcttggtac cgagctcgga
      901 tccatggctt tccgcaggag gacgaaaagt tacccgctct tcagccagga gttcgtcatc
      961 cacaaccatg cggacatcgg cttctgcctg gtgctctgcg tcctcatcgg gcttatgttc
     1021 gaggtcacag ccaagactgc ctttctattt attttacctc agtataacat tagcgtgcct
     1081 acagcagaca gtgagaccgt gcactaccac tatggcccta aggacctggt cacaatcttg
     1141 ttctacatct tcatcaccat catcttgcat gctgtggttc aggagtacat tttagataaa
     1201 atcagcaaac ggcttcatct ctccaaagtc aaacacagca agttcaatga atctggacag
     1261 ctggtcgtct ttcatttcac ctcggtgatt tggtgcttct acgtggtggt gacggaagga
     1321 tacttaacaa acccaagaag cctctgggaa gactacccgc atgtgcacct ccccttccag
     1381 gtgaagtttt tctacctatg ccagctggcc tactggctgc acgcacttcc tgagctatac
     1441 ttccagaagg tacggaagga ggaaattccc cgccagctcc agtatatttg cctgtacctg
     1501 gggcatatag ctggagcata cctcttaaac ctgagccgcc tgggcctgat cttgctgctg
     1561 ctgcagtact caactgagtt cctcttccac acggctagac tcttctactt tgcagatgaa
     1621 aacaacgaga aactgttcag tgcctgggct gctgtttttg gggttacccg cctcttcatc
     1681 ctcacccttg ccgtgctggc cattggcttt ggactggctc gcatggaaaa ccaggcattt
     1741 gatcccgaga aagggaactt caacactttg ttttgcaggc tctgcgtgct gctgctggtg
     1801 tgtgccgccc aggcctggct catgtggcgc ttcatccact cccagctgcg gcactggcgg
     1861 gaatactgga atgagcagag tgcaaagcgg agagtcccag ccacacccag actaccagcc
     1921 aggctcatca agagggaatc tggttaccat gaaaatggag tggtgaaggc agagaacgga
     1981 acctccccac ggactaagaa actcaagtct cccgaattcg ccgccaccat ggcggccgcc
     2041 agctggagcc accctcagtt cgagaaggga ggaggaagcg gcggaggcag cggaggagga
     2101 agctggagcc acccgcagtt cgagaaatga ctcgagcatg catctagagg ctggagccac
     2161 ccgcagttcg agaaatgact cgagcatgca tctagagggc cctattctat agtgtcacct
     2221 aaatgctaga gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     2281 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     2341 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     2401 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     2461 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     2521 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2581 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2641 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2701 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2761 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2821 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2881 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2941 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3001 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3061 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     3121 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     3181 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     3241 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     3301 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     3361 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3421 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3481 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     3541 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3601 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3661 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3721 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3781 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     3841 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     3901 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     3961 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4021 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4081 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4141 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     4201 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4261 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4321 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     4381 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     4441 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     4501 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4561 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4621 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4681 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4741 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4801 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     4861 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     4921 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     4981 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5041 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5101 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5161 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5221 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5281 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     5341 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5401 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5461 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5521 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5581 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5641 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5701 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5761 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5821 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5881 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5941 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6001 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6061 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6121 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6181 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6241 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6301 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6361 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6421 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6481 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6541 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6601 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     6661 cg
//
