LOCUS       sequence-242546-        9414 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector for expression of Nedd8 attachment mutant K705R 
            CUL4A-P2A-Puro casette from a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9414)
  AUTHORS   Patil A, Manzano M, Gottwein E
  TITLE     Genome-wide CRISPR screens reveal genetic mediators of cereblon 
            modulator toxicity in primary effusion lymphoma.
  JOURNAL   Blood Adv. 2019 Jul 23;3(14):2105-2117. doi: 
            10.1182/bloodadvances.2019031732.
  PUBMED    31300418
REFERENCE   2  (bases 1 to 9414)
  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: 
            "10.1182/bloodadvances"; journalName: "Blood Adv"; date: 
            "2019-07-23- 23"; volume: "3"; issue: "14"; pages: "2105-2117"
COMMENT     Sequence Label: pLC-CUL4A K705R-P2A-Puro
FEATURES             Location/Qualifiers
     source          1..9414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2287..2343
                     /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             2344..2940
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     2837..2857
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(2977..3010)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3066..3654
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3119..3139)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3537..3548)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3657..3673)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3658..3674)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             4183..4363
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(4425..5013)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4505..4524)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5184..5975,5976..6044))
                     /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     5807..5826
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6045..6149)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6224..6243)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6415..6794
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6796..6994
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             7012..7192
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7239..7364
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7861..8094
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8279..8323
                     /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    8621..8738
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        8863..9166
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        9167..9370
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      join(9409..9414,1..4)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 atggcggacg aggccccgcg gaagggcagc ttctcggcgc tcgtgggccg caccaacggc
       61 ctcaccaagc ccgcggccct ggccgccgcg cccgccaagc cggggggcgc gggcggctcc
      121 aagaagctgg tcatcaagaa cttccgagac agacctcggc tgcccgacaa ctacacgcag
      181 gacacgtggc ggaagctgca cgaggcggtg cgggccgtgc agagcagcac ctccatcagg
      241 tacaacctcg aggagctcta ccaggctgtg gaaaatctct gttctcacaa agtctcccca
      301 atgctctaca agcaactgcg tcaggcctgt gaagaccacg tccaggcaca gatccttccg
      361 tttagagaag actcactaga tagtgtttta tttttaaaga agattaacac gtgctggcag
      421 gaccactgca gacaaatgat catgatcaga agcatcttcc tgttcttgga ccgcacctat
      481 gtgctgcaga actccacgct gccctccatc tgggatatgg gattagaact gtttagaacc
      541 catattatta gtgataaaat ggttcagagt aaaaccattg atggaatcct actgctgatc
      601 gagcgcgaga ggagcggcga ggccgtggac cggagcctgt tgcggagcct cctgggcatg
      661 ctgtctgacc tgcaggtgta taaagattca tttgaactga aatttttgga agagactaat
      721 tgcttatatg ctgccgaagg ccaaaggtta atgcaggaaa gagaggttcc agaatatctt
      781 aaccatgtaa gtaaacgctt agaggaagag ggagacagag taatcactta cttggaccac
      841 agcacacaga aaccactgat tgcttgtgtg gagaaacagc tattaggaga acatttaaca
      901 gcaattctgc agaaagggct cgaccactta ctggatgaga acagagtgcc ggacctcgca
      961 cagatgtacc agctgttcag ccgggtgagg ggcgggcagc aggcgctgct gcagcactgg
     1021 agcgagtaca tcaagacttt tggaacagcg atcgtaatca atcctgagaa agacaaagac
     1081 atggtccaag acctgttgga cttcaaggac aaggtggacc acgtgatcga ggtctgcttc
     1141 cagaagaatg agcggttcgt caacctgatg aaggagtcct ttgagacgtt catcaacaag
     1201 agacccaaca agcctgcaga actgatcgca aagcatgtgg attcaaagtt aagagcaggc
     1261 aacaaagaag ccacagacga ggagctggag cggacgttgg acaagatcat gatcctgttc
     1321 aggtttatcc acggtaaaga tgtctttgaa gcattttata aaaaagattt ggcaaaaaga
     1381 ctccttgttg ggaaaagtgc ctcagtcgat gctgaaaagt ctatgttgtc aaagctcaag
     1441 catgagtgcg gtgcagcctt caccagcaag ctggaaggca tgttcaagga catggagctt
     1501 tcgaaggaca tcatggttca tttcaagcag catatgcaga atcagagtga ctcaggccct
     1561 atagacctca cagtgaacat actcacaatg ggctactggc caacatacac gcccatggaa
     1621 gtgcacttaa ccccagaaat gattaaactt caggaagtat ttaaggcatt ttatcttgga
     1681 aagcacagtg gtcgaaaact tcagtggcaa actactttgg gacatgctgt tttaaaagcg
     1741 gagtttaaag aagggaagaa ggaattccag gtgtccctct tccagacact ggtgctcctc
     1801 atgttcaacg agggagatgg cttcagcttt gaggagataa aaatggccac ggggatagag
     1861 gatagtgaat tgcgcagaac gctgcagtcc ctggcctgtg gcaaagcacg tgtgctgatt
     1921 aaaagtccca aaggaaagga agtggaagat ggagacaagt tcatttttaa tggagagttc
     1981 aagcacaagt tgtttagaat aaagatcaat caaattcaga tgaaggaaac tgttgaggaa
     2041 caggttagca ccactgagag agtgtttcag gatagacaat atcagattga tgctgctatc
     2101 gtcagaataa tgcggatgag aaagactctt ggtcataatc ttctagtttc tgaattatat
     2161 aatcagctga aatttccagt aaagcctgga gatttgaaaa agagaattga atctctgata
     2221 gacagagact atatggagag agacaaagac aatccgaatc agtaccacta cgtggccgga
     2281 tccggcgcaa caaacttctc tctgctgaaa caagccggag atgtcgaaga gaatcctgga
     2341 ccgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     2401 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     2461 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     2521 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     2581 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     2641 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     2701 cccgcgtggt tcctggccac cgtcggagtc tcgcccgacc accagggcaa gggtctgggc
     2761 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     2821 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     2881 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     2941 aatatgcggc cgccagagaa ttcgtcgagg gacctaataa cttcgtatag catacattat
     3001 acgaagttat acatgtttaa gggttccggt tccactaggt acaattcgat atcaagctta
     3061 tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     3121 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     3181 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     3241 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     3301 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     3361 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     3421 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     3481 tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
     3541 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
     3601 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat
     3661 accgtcgacc tcgatcgaga cctagaaaaa catggagcaa tcacaagtag caatacagca
     3721 gctaccaatg ctgattgtgc ctggctagaa gcacaagagg aggaggaggt gggttttcca
     3781 gtcacacctc aggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     3841 tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatatc
     3901 cttgatctgt ggatctacca cacacaaggc tacttccctg attggcagaa ctacacacca
     3961 gggccaggga tcagatatcc actgaccttt ggatggtgct acaagctagt accagttgag
     4021 caagagaagg tagaagaagc caatgaagga gagaacaccc gcttgttaca ccctgtgagc
     4081 ctgcatggga tggatgaccc ggagagagaa gtattagagt ggaggtttga cagccgccta
     4141 gcatttcatc acatggcccg agagctgcat ccggactgta ctgggtctct ctggttagac
     4201 cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa
     4261 agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag
     4321 agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagcatgtg agcaaaaggc
     4381 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4441 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4501 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4561 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4621 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4681 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4741 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4801 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4861 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4921 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4981 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     5041 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     5101 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5161 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5221 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5281 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5341 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5401 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5461 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5521 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5581 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5641 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5701 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5761 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5821 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5881 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5941 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     6001 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     6061 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6121 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     6181 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
     6241 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
     6301 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
     6361 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
     6421 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     6481 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     6541 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     6601 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     6661 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     6721 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     6781 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
     6841 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
     6901 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
     6961 gtaggcgtgt acggtgggag gtctatataa gcagcgcgtt ttgcctgtac tgggtctctc
     7021 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
     7081 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
     7141 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
     7201 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
     7261 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
     7321 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
     7381 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
     7441 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
     7501 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
     7561 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
     7621 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
     7681 gtaagaccac cgcacagcaa gcggccggcc gctgatcttc agacctggag gaggagatat
     7741 gagggacaat tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg
     7801 agtagcaccc accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat
     7861 aggagctttg ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat
     7921 gacgctgacg gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt
     7981 gctgagggct attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca
     8041 gctccaggca agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat
     8101 ttggggttgc tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag
     8161 taataaatct ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat
     8221 taacaattac acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa
     8281 gaatgaacaa gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat
     8341 aacaaattgg ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt
     8401 aagaatagtt tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt
     8461 atcgtttcag acccacctcc caaccccgag gggacccgac aggcccgaag gaatagaaga
     8521 agaaggtgga gagagagaca gagacagatc cattcgatta gtgaacggat cggcactgcg
     8581 tgcgccaatt ctgcagacaa atggcagtat tcatccacaa ttttaaaaga aaagggggga
     8641 ttggggggta cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta
     8701 aagaattaca aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca
     8761 gagatccagt ttggttagta ccgggcccga gagttaatta attagttatt aatagtaatc
     8821 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     8881 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     8941 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     9001 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     9061 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     9121 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     9181 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     9241 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     9301 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     9361 aagcagagct ggtttagtga accgtcagat ccgctagcgc taccggtcgc cacc
//
