LOCUS       sequence-340221-        6127 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral vector for the constitutive, near-physiological 
            expression of 2xFLAG-2xSTREP-tagged cyclin D2 and a puromycin 
            resistance marker in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6127)
  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 6127)
  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-CCND2-Puro
FEATURES             Location/Qualifiers
     source          1..6127
                     /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"
     primer_bind     complement(2457..2477)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2462..2791
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2642..2777
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2801..3400
                     /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(2801..2820)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3297..3317
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3621..4090
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(4220..4237)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4391..4979)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4471..4490)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5139..5930,5931..5999))
                     /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     5762..5781
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6000..6104)
                     /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 ctagcagcga gctgctgtgc cacgaggtgg acccggtccg cagggccgtg
     1621 cgggaccgca acctgctccg agacgaccgc gtcctgcaga acctgctcac catcgaggag
     1681 cgctaccttc cgcagtgctc ctacttcaag tgcgtgcaga aggacatcca accctacatg
     1741 cgcagaatgg tggccacctg gatgctggag gtctgtgagg aacagaagtg cgaagaagag
     1801 gtcttccctc tggccatgaa ttacctggac cgtttcttgg ctggggtccc gactccgaag
     1861 tcccatctgc aactcctggg tgctgtctgc atgttcctgg cctccaaact caaagagacc
     1921 agcccgctga ccgcggagaa gctgtgcatt tacaccgaca actccatcaa gcctcaggag
     1981 ctgctggagt gggaactggt ggtgctgggg aagttgaagt ggaacctggc agctgtcact
     2041 cctcatgact tcattgagca catcttgcgc aagctgcccc agcagcggga gaagctgtct
     2101 ctgatccgca agcatgctca gaccttcatt gctctgtgtg ccacggactt taagtttgcc
     2161 atgtacccac cgtcgatgat cgcaactgga agtgtgggag cagccatctg tgggctccag
     2221 caggatgagg aagtgagctc gctcacttgt gatgccctga ctgagctgct ggctaagatc
     2281 accaacacag acgtggattg tctcaaagct tgccaggagc agattgaggc ggtgctcctc
     2341 aatagcctgc agcagtaccg tcaggaccaa cgtgacggat ccaagtcgga ggatgaactg
     2401 gaccaagcca gcacccctac agacgtgcgg gatatcgacc tgtgactcga ccctgtggaa
     2461 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     2521 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     2581 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     2641 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     2701 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     2761 aggctttttt ggaggcctag gcttttgcaa aaagcttacc atgaccgagt acaagcccac
     2821 ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt
     2881 cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac
     2941 cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc
     3001 ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt
     3061 gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca
     3121 acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac
     3181 cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg
     3241 agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa
     3301 cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg
     3361 accgcgcacc tggtgcatga cccgcaagcc cggtgcctga cgcccgcccc acgacccgca
     3421 gcgcccgacc gaaaggagcg cacgacccca tgcatcgata aaataaaaga ttttatttag
     3481 tctccagaaa aaggggggaa tgaaagaccc cacctgtagg tttggcaagc tagcttaagt
     3541 aacgccattt tgcaaggcat ggaaaaatac ataactgaga atagagaagt tcagatcaag
     3601 gtcaggaaca gatggaacag ctgaatatgg gccaaacagg atatctgtgg taagcagttc
     3661 ctgccccggc tcagggccaa gaacagatgg aacagctgaa tatgggccaa acaggatatc
     3721 tgtggtaagc agttcctgcc ccggctcagg gccaagaaca gatggtcccc agatgcggtc
     3781 cagccctcag cagtttctag agaaccatca gatgtttcca gggtgcccca aggacctgaa
     3841 atgaccctgt gccttatttg aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc
     3901 ttctgctccc cgagctcaat aaaagagccc acaacccctc actcggggcg ccagtcctcc
     3961 gattgactga gtcgcccggg tacccgtgta tccaataaac cctcttgcag ttgcatccga
     4021 cttgtggtct cgctgttcct tgggagggtc tcctctgagt gattgactac ccgtcagcgg
     4081 gggtctttca catgcagcat gtatcaaaat taatttggtt ttttttctta agtatttaca
     4141 ttaaatggcc atagttgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     4201 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     4261 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     4321 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     4381 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     4441 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     4501 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     4561 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4621 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4681 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4741 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4801 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     4861 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4921 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4981 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     5041 ggattttggt catgagatta tcaaaaagga tcttcaccta gatccttttg cggccgcaaa
     5101 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     5161 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     5221 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     5281 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     5341 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     5401 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     5461 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     5521 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5581 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     5641 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     5701 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     5761 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     5821 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     5881 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     5941 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     6001 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     6061 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     6121 gtgccac
//
