LOCUS       sequence-340219-        6145 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral vector for the constitutive, near-physiological 
            expression of 2xFLAG-2xSTREP-tagged cyclin D1(T286A) and a puromycin
            resistance marker in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6145)
  AUTHORS   Simoneschi D, Rona G, Zhou N, Jeong YT, Jiang S, Milletti G, Arbini 
            AA, O'Sullivan A, Wang AA, Nithikasem S, Keegan S, Siu Y, 
            Cianfanelli V, Maiani E, Nazio F, Cecconi F, Boccalatte F, Fenyo D, 
            Jones DR, Busino L, Pagano M
  TITLE     CRL4(AMBRA1) is a master regulator of D-type cyclins.
  JOURNAL   Nature. 2021 Apr;592(7856):789-793. doi: 10.1038/s41586-021-03445-y.
            Epub 2021 Apr 14.
  PUBMED    33854235
REFERENCE   2  (bases 1 to 6145)
  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: 
            "10.1038/s41586-021-03445-y"; journalName: "Nature"; date: 
            "2021-04"; volume: "592"; issue: "7856"; pages: "789-793"
COMMENT     Sequence Label: pBABE-2xFLAG-2xSTREP-CCND1(T286A)-Puro
FEATURES             Location/Qualifiers
     source          1..6145
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             8..477
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    542..741
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             942..1358
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1288..1310
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1326..1342
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             1399..1422
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1438..1461
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1477..1500
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1537..1560
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     regulatory      2177..2186
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(2475..2495)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2480..2809
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2660..2795
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2819..3418
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2819..2838)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3315..3335
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3639..4108
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(4238..4255)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4409..4997)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4489..4508)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5157..5948,5949..6017))
                     /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     5780..5799
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6018..6122)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgcagcctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca
       61 gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt ggtaagcagt
      121 tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag ccctcagcag
      181 tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg accctgtgcc
      241 ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga
      301 gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat tgactgagtc
      361 gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt gtggtctcgc
      421 tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg tctttcattt
      481 gggggctcgt ccgggatcgg gagacccctg cccagggacc accgacccac caccgggagg
      541 caagctggcc agcaacttat ctgtgtctgt ccgattgtct agtgtctatg actgatttta
      601 tgcgcctgcg tcggtactag ttagctaact agctctgtat ctggcggacc cgtggtggaa
      661 ctgacgagtt ctgaacaccc ggccgcaacc ctgggagacg tcccagggac tttgggggcc
      721 gtttttgtgg cccgacctga ggaagggagt cgatgtggaa tccgaccccg tcaggatatg
      781 tggttctggt aggagacgag aacctaaaac agttcccgcc tccgtctgaa tttttgcttt
      841 cggtttggaa ccgaagccgc gcgtcttgtc tgctgcagcg ctgcagcatc gttctgtgtt
      901 gtctctgtct gactgtgttt ctgtatttgt ctgaaaatta gggccagact gttaccactc
      961 ccttaagttt gaccttaggt cactggaaag atgtcgagcg gatcgctcac aaccagtcgg
     1021 tagatgtcaa gaagagacgt tgggttacct tctgctctgc agaatggcca acctttaacg
     1081 tcggatggcc gcgagacggc acctttaacc gagacctcat cacccaggtt aagatcaagg
     1141 tcttttcacc tggcccgcat ggacacccag accaggtccc ctacatcgtg acctgggaag
     1201 ccttggcttt tgacccccct ccctgggtca agccctttgt acaccctaag cctccgcctc
     1261 ctcttcctcc atccgccccg tctctccccc ttgaacctcc tcgttcgacc ccgcctcgat
     1321 cctcccttta tccagccctc actccttctc taggcgccgg ccggatccca gtgtggtggt
     1381 acgtaggaat tcagcatgga ctacaaggac gacgatgaca aggtaccagg atctggggat
     1441 tataaagatg atgatgataa agggtcggcc gccagctgga gccaccctca gttcgagaag
     1501 ggaggaggaa gcggcggagg cagcggagga ggaagctgga gccacccgca gttcgagaaa
     1561 ggagctggat ccagcagcga acaccagctc ctgtgctgcg aagtggaaac catccgccgc
     1621 gcgtaccccg atgccaacct cctcaacgac cgggtgctgc gggccatgct gaaggcggag
     1681 gagacctgcg cgccctcggt gtcctacttc aaatgtgtgc agaaggaggt cctgccgtcc
     1741 atgcggaaga tcgtcgccac ctggatgctg gaggtctgcg aggaacagaa gtgcgaggag
     1801 gaggtcttcc cgctggccat gaactacctg gaccgcttcc tgtcgctgga gcccgtgaaa
     1861 aagagccgcc tgcagctgct gggggccact tgcatgttcg tggcctctaa gatgaaggag
     1921 accatccccc tgacggccga gaagctgtgc atctacaccg acaactccat ccggcccgag
     1981 gagctgctgc aaatggagct gctcctggtg aacaagctca agtggaacct ggccgcaatg
     2041 accccgcacg atttcattga acacttcctc tccaaaatgc cagaggcgga ggagaacaaa
     2101 cagatcatcc gcaaacacgc gcagaccttc gttgccctct gtgccacaga tgtgaagttc
     2161 atttccaatc cgccctccat ggtggcagcg gggagcgtgg tggccgcagt gcaaggcctg
     2221 aacctgagga gccccaacaa cttcctgtcc tactaccgcc tcacacgctt cctctccaga
     2281 gtgatcaagt gtgacccaga ctgcctccgg gcctgccagg agcagatcga agccctgctg
     2341 gagtcaagcc tgcgccaggc ccagcagaac atggacccca aggccgccga ggaggaggaa
     2401 gaggaggagg aggaggtgga cctggcttgc gcacccaccg acgtgcggga cgtggacatc
     2461 tgactcgacc ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     2521 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     2581 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     2641 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     2701 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     2761 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agcttaccat
     2821 gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg
     2881 caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg
     2941 ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat
     3001 cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag
     3061 cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc
     3121 ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc
     3181 cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag
     3241 cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga
     3301 gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga
     3361 cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg
     3421 cccgccccac gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcgataaa
     3481 ataaaagatt ttatttagtc tccagaaaaa ggggggaatg aaagacccca cctgtaggtt
     3541 tggcaagcta gcttaagtaa cgccattttg caaggcatgg aaaaatacat aactgagaat
     3601 agagaagttc agatcaaggt caggaacaga tggaacagct gaatatgggc caaacaggat
     3661 atctgtggta agcagttcct gccccggctc agggccaaga acagatggaa cagctgaata
     3721 tgggccaaac aggatatctg tggtaagcag ttcctgcccc ggctcagggc caagaacaga
     3781 tggtccccag atgcggtcca gccctcagca gtttctagag aaccatcaga tgtttccagg
     3841 gtgccccaag gacctgaaat gaccctgtgc cttatttgaa ctaaccaatc agttcgcttc
     3901 tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa aagagcccac aacccctcac
     3961 tcggggcgcc agtcctccga ttgactgagt cgcccgggta cccgtgtatc caataaaccc
     4021 tcttgcagtt gcatccgact tgtggtctcg ctgttccttg ggagggtctc ctctgagtga
     4081 ttgactaccc gtcagcgggg gtctttcaca tgcagcatgt atcaaaatta atttggtttt
     4141 ttttcttaag tatttacatt aaatggccat agttgcatta atgaatcggc caacgcgcgg
     4201 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     4261 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     4321 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     4381 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     4441 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     4501 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     4561 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     4621 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     4681 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     4741 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     4801 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     4861 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     4921 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     4981 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5041 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     5101 tccttttgcg gccgcaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     5161 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     5221 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     5281 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     5341 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     5401 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     5461 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     5521 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     5581 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     5641 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     5701 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     5761 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     5821 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     5881 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     5941 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     6001 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     6061 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     6121 cgcacatttc cccgaaaagt gccac
//
