LOCUS       sequence-361671-        6898 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human dihydrofolate reductase (DHFR) with C-terminal HiBiT
            tag in mammalian cells. This retroviral vector has an IRES-Lyt-2 
            downstream of the cloning sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6898)
  AUTHORS   Rana S, Dranchak P, Dahlin JL, Lamy L, Li W, Oliphant E, Shrimp JH, 
            Rajacharya GH, Tharakan R, Holland DO, Whitten AS, Wilson KM, Singh 
            PK, Durum SK, Tao D, Rai G, Inglese J
  TITLE     Methotrexate-based PROTACs as DHFR-specific chemical probes.
  JOURNAL   Cell Chem Biol. 2024 Feb 15;31(2):221-233.e14. doi: 
            10.1016/j.chembiol.2023.09.014. Epub 2023 Oct 23.
  PUBMED    37875111
REFERENCE   2  (bases 1 to 6898)
  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.chembiol.2023.09.014"; journalName: "Cell Chem Biol"; 
            date: "2024-02-15- 15"; volume: "31"; issue: "2"; pages: 
            "221-233.e14"
COMMENT     Sequence Label: pBMN DHFR-HiBiT / Lyt2
FEATURES             Location/Qualifiers
     source          1..6898
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    join(2..576,577..579,580..588)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(169..186)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     396..415
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             589..1254
                     /codon_start=1
                     /product="mouse lymphocyte antigen CD8 alpha chain"
                     /label=mCD8
                     /translation="MASPLTRFLSLNLLLLGESIILGSGEAKPQAPELRIFPKKMDAEL
                     GQKVDLVCEVLGSVSQGCSWLFQNSSSKLPQPTFVVYMASSHNKITWDEKLNSSKLFSA
                     MRDTNNKYVLTLNKFSKENEGYYFCSVISNSVMYFSSVVPVLQKVNSTTTKPVLRTPSP
                     VHPTGTSQPQRPEDCRPRGSVKGTGLDFACDIYIWAPLAGICVALLLSLIITLICYHSR
                     "
     LTR             1475..1944
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(2074..2091)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2245..2833)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2325..2344)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(2957..3025,3026..3817)
                     /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(3175..3194)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     LTR             4499..4968
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    5033..5390
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             5455..5871
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     5801..5823
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     5839..5855
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    5881..6255
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     6193..6209
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     regulatory      6264..6273
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 ccccctctcc ctcccccccc cctaacgtta ctggccgaag ccgcttggaa taaggccggt
       61 gtgcgtttgt ctatatgtta ttttccacca tattgccgtc ttttggcaat gtgagggccc
      121 ggaaacctgg ccctgtcttc ttgacgagca ttcctagggg tctttcccct ctcgccaaag
      181 gaatgcaagg tctgttgaat gtcgtgaagg aagcagttcc tctggaagct tcttgaagac
      241 aaacaacgtc tgtagcgacc ctttgcaggc agcggaaccc cccacctggc gacaggtgcc
      301 tctgcggcca aaagccacgt gtataagata cacctgcaaa ggcggcacaa ccccagtgcc
      361 acgttgtgag ttggatagtt gtggaaagag tcaaatggct ctcctcaagc gtattcaaca
      421 aggggctgaa ggatgcccag aaggtacccc attgtatggg atctgatctg gggcctcggt
      481 gcacatgctt tacatgtgtt tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg
      541 gggacgtggt tttcctttga aaaacacgat gataatatgg ccacaaccat ggcctcaccg
      601 ttgacccgct ttctgtcgct gaacctgctg ctgctgggtg agtcgattat cctggggagt
      661 ggagaagcta agccacaggc acccgaactc cgaatctttc caaagaaaat ggacgccgaa
      721 cttggtcaga aggtggacct ggtatgtgaa gtgttggggt ccgtttcgca aggatgctct
      781 tggctcttcc agaactccag ctccaaactc ccccagccca ccttcgttgt ctatatggct
      841 tcatcccaca acaagataac gtgggacgag aagctgaatt cgtcgaaact gttttctgcc
      901 atgagggaca cgaataataa gtacgttctc accctgaaca agttcagcaa ggaaaacgaa
      961 ggctactatt tctgctcagt catcagcaac tcggtgatgt acttcagttc tgtcgtgcca
     1021 gtccttcaga aagtgaactc tactactacc aagccagtgc tgcgaactcc ctcacctgtg
     1081 caccctaccg ggacatctca gccccagaga ccagaagatt gtcggccccg tggctcagtg
     1141 aaggggaccg gattggactt cgcctgtgat atttacatct gggcaccctt ggccggaatc
     1201 tgcgtggccc ttctgctgtc cttgatcatc actctcatct gctaccacag ccgctagtca
     1261 gacaggaaga tcgggccgcg gggatcatca agcttatcga taccgtcgac gataaaataa
     1321 aagattttat ttagtctcca gaaaaagggg ggaatgaaag accccacctg taggtttggc
     1381 aagctagctt aagtaacgcc attttgcaag gcatggaaaa atacataact gagaatagag
     1441 aagttcagat caaggtcagg aacagatgga acagctgaat atgggccaaa caggatatct
     1501 gtggtaagca gttcctgccc cggctcaggg ccaagaacag atggaacagc tgaatatggg
     1561 ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag aacagatggt
     1621 ccccagatgc ggtccagccc tcagcagttt ctagagaacc atcagatgtt tccagggtgc
     1681 cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa ccaatcagtt cgcttctcgc
     1741 ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc cctcactcgg
     1801 ggcgccagtc ctccgattga ctgagtcgcc cgggtacccg tgtatccaat aaaccctctt
     1861 gcagttgcat ccgacttgtg gtctcgctgt tccttgggag ggtctcctct gagtgattga
     1921 ctacccgtca gcgggggtct ttcacatgca gcatgtatca aaattaattt ggtttttttt
     1981 cttaagtatt tacattaaat ggccatagtt gcattaatga atcggccaac gcgcggggag
     2041 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     2101 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     2161 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     2221 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     2281 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     2341 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     2401 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     2461 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     2521 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     2581 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     2641 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     2701 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     2761 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     2821 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     2881 aaactcacgt taagggattt tggtcatgag acaataaccc tgataaatgc ttcaataata
     2941 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     3001 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     3061 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     3121 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     3181 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     3241 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     3301 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     3361 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     3421 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     3481 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     3541 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     3601 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     3661 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     3721 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     3781 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     3841 tatactttag attgatttgc ggccggccgc aaaaggatct aggtgaagat cctttttgat
     3901 aatctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     3961 cgcacatttc cccgaaaagt gccaccagct ttgctcttag gagtttccta atacatccca
     4021 aactcaaata tataaagcat ttgacttgtt ctatgcccta gggggcgggg ggaagctaag
     4081 ccagcttttt ttaacattta aaatgttaat tccattttaa atgcacagat gtttttattt
     4141 cataagggtt tcaatgtgca tgaatgctgc aatattcctg ttaccaaagc tagtataaat
     4201 aaaaatagat aaacgtggaa attacttaga gtttctgtca ttaacgtttc cttcctcagt
     4261 tgacaacata aatgcgctgc tgagaagcca gtttgcatct gtcaggatca atttcccatt
     4321 atgccagtca tattaattac tagtcaatta gttgattttt atttttgaca tatacatgtg
     4381 aaagacccca cctgtaggtt tggcaagcta gcttaagtaa cgccattttg caaggcatgg
     4441 aaaaatacat aactgagaat agaaaagttc agatcaaggt caggaacaga tggaacagct
     4501 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
     4561 acagatggaa cagctgaata tgggccaaac aggatatctg tggtaagcag ttcctgcccc
     4621 ggctcagggc caagaacaga tggtccccag atgcggtcca gccctcagca gtttctagag
     4681 aaccatcaga tgtttccagg gtgccccaag gacctgaaat gaccctgtgc cttatttgaa
     4741 ctaaccaatc agttcgcttc tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa
     4801 aagagcccac aacccctcac tcggcgcgcc agtcctccga ttgactgagt cgcccgggta
     4861 cccgtgtatc caataaaccc tcttgcagtt gcatccgact tgtggtctcg ctgttccttg
     4921 ggagggtctc ctctgagtga ttgactaccc gtcagcgggg gtctttcatt tgggggctcg
     4981 tccgggatcg ggagacccct gcccagggac caccgaccca ccaccgggag gtaagctggc
     5041 cagcaactta tctgtgtctg tccgattgtc tagtgtctat gactgatttt atgcgcctgc
     5101 gtcggtacta gttagctaac tagctctgta tctggcggac ccgtggtgga actgacgagt
     5161 tcggaacacc cggccgcaac cctgggagac gtcccaggga cttcgggggc cgtttttgtg
     5221 gcccgacctg agtccaaaaa tcccgatcgt tttggactct ttggtgcacc ccccttagag
     5281 gagggatatg tggttctggt aggagacgag aacctaaaac agttcccgcc tccgtctgaa
     5341 tttttgcttt cggtttggga ccgaagccgc gccgcgcgtc ttgtctgctg cagcatcgtt
     5401 ctgtgttgtc tctgtctgac tgtgtttctg tatttgtctg aaaatatggg cccgggccag
     5461 actgttacca ctcccttaag tttgacctta ggtcactgga aagatgtcga gcggatcgct
     5521 cacaaccagt cggtagatgt caagaagaga cgttgggtta ccttctgctc tgcagaatgg
     5581 ccaaccttta acgtcggatg gccgcgagac ggcaccttta accgagacct catcacccag
     5641 gttaagatca aggtcttttc acctggcccg catggacacc cagaccaggt cccctacatc
     5701 gtgacctggg aagccttggc ttttgacccc cctccctggg tcaagccctt tgtacaccct
     5761 aagcctccgc ctcctcttcc tccatccgcc ccgtctctcc cccttgaacc tcctcgttcg
     5821 accccgcctc gatcctccct ttatccagcc ctcactcctt ctctaggcgc ccccatatgg
     5881 ccatatgaga tcttatatgg ggcacccccg ccccttgtaa acttccctga ccctgacatg
     5941 acaagagtta ctaacagccc ctctctccaa gctcacttac aggctctcta cttagtccag
     6001 cacgaagtct ggagacctct ggcggcagcc taccaagaac aactggaccg accggtggta
     6061 cctcaccctt accgagtcgg cgacacagtg tgggtccgcc gacaccagac taagaaccta
     6121 gaacctcgct ggaaaggacc ttacacagtc ctgctgacca cccccaccgc cctcaaagta
     6181 gacggcatcg cagcttggat acacgccgcc cacgtgaagg ctgccgaccc cgggggtgga
     6241 ccatcctcta gactgccgga tccgccacca tggttggttc gctaaactgc atcgtcgctg
     6301 tgtcccagaa catgggcatc ggcaagaacg gggacctgcc ctggccaccg ctcaggaatg
     6361 aattcagata tttccagaga atgaccacaa cctcttcagt agaaggtaaa cagaatctgg
     6421 tgattatggg taagaagacc tggttctcca ttcctgagaa gaatcgacct ttaaagggta
     6481 gaattaattt agttctcagc agagaactca aggaacctcc acaaggagct cattttcttt
     6541 ccagaagtct agatgatgcc ttaaaactta ctgaacaacc agaattagca aataaagtag
     6601 acatggtctg gatagttggt ggcagttctg tttataagga agccatgaat cacccaggcc
     6661 atcttaaact atttgtgaca aggatcatgc aagactttga aagtgacacg ttttttccag
     6721 aaattgattt ggagaaatat aaacttctgc cagaataccc aggtgttctc tctgatgtcc
     6781 aggaggagaa aggcattaag tacaaatttg aagtatatga gaagaatgat gtgagcggct
     6841 ggcggctgtt caagaagatt agctaataag cggccgctac gtaaattccg cccccccc
//
