LOCUS       sequence-448496-        6554 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of wild type human Rb.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6554)
  AUTHORS   Narasimha AM, Kaulich M, Shapiro GS, Choi YJ, Sicinski P, Dowdy SF
  TITLE     Cyclin D activates the Rb tumor suppressor by mono-phosphorylation.
  JOURNAL   Elife. 2014 May 29:e02872. doi: 10.7554/eLife.02872.
  PUBMED    24876129
REFERENCE   2  (bases 1 to 6554)
  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: "Elife. 2014
            May 29:e02872. doi: 10.7554/eLife.02872."
COMMENT     Sequence Label: pCMV HA hRB-wt
FEATURES             Location/Qualifiers
     source          1..6554
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     30..52
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     44..61
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     45..61
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        88..391
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        392..595
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     546..566
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     592..616
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          733..829
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             893..919
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    3754..3888
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3778..3797)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3832..3851
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3999..4015)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3999..4015)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4012..4034)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4023..4039
                     /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(4047..4053,4054..4071,4072..4077))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4092..4113
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4230..4247)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4401..4989)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4481..4500)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5160..5951,5952..6020))
                     /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     5783..5802
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6021..6125)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6193..6211
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6249..6271)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6371..6390
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag
       61 tgagttcgag cttgcatgcc tgcaggtcgt tacataactt acggtaaatg gcccgcctgg
      121 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
      181 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
      241 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
      301 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
      361 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
      421 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
      481 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
      541 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctcgttt
      601 agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc atagaagaca
      661 ccgggaccga tccagcctcc ggactctaga ggatccggta ctagaggaac tgaaaaacca
      721 gaaagttaac tggtaagttt agtctttttg tcttttattt caggtcccgg atccggtggt
      781 ggtgcaaatc aaagaactgc tcctcagtgg atgttgcctt tacttctagg cctgtacgga
      841 agtgttactt ctgctctaaa agctgcggaa ttgtacccgc gggcccacca tgtacccata
      901 cgatgttcca gattacgctc ttatggccat ggaggcccga attcggtcga ccccgcccaa
      961 aaccccccga aaaacggccg ccaccgccgc cgctgccgcc gcggaacccc cggcaccgcc
     1021 gccgccgccc cctcctgagg aggacccaga gcaggacagc ggcccggagg acctgcctct
     1081 cgtcaggctt gagtttgaag aaacagaaga acctgatttt actgcattat gtcagaaatt
     1141 aaagatacca gatcatgtca gagagagagc ttggttaact tgggagaaag tttcatctgt
     1201 ggatggagta ttgggaggtt atattcaaaa gaaaaaggaa ctgtggggaa tctgtatctt
     1261 tattgcagca gttgacctag atgagatgtc gttcactttt actgagctac agaaaaacat
     1321 agaaatcagt gtccataaat tctttaactt actaaaagaa attgatacca gtaccaaagt
     1381 tgataatgct atgtcaagac tgttgaagaa gtatgatgta ttgtttgcac tcttcagcaa
     1441 attggaaagg acatgtgaac ttatatattt gacacaaccc agcagttcga tatctactga
     1501 aataaattct gcattggtgc taaaagtttc ttggatcaca tttttattag ctaaagggga
     1561 agtattacaa atggaagatg atctggtgat ttcatttcag ttaatgctat gtgtccttga
     1621 ctattttatt aaactctcac ctcccatgtt gctcaaagaa ccatataaaa cagctgttat
     1681 acccattaat ggttcacctc gaacacccag gcgaggtcag aacaggagtg cacggatagc
     1741 aaaacaacta gaaaatgata caagaattat tgaagttctc tgtaaagaac atgaatgtaa
     1801 tatagatgag gtgaaaaatg tttatttcaa aaattttata ccttttatga attctcttgg
     1861 acttgtaaca tctaatggac ttccagaggt tgaaaatctt tctaaacgat acgaagaaat
     1921 ttatcttaaa aataaagatc tagatgcaag attatttttg gatcatgata aaactcttca
     1981 gactgattct atagacagtt ttgaaacaca gagaacacca cgaaaaagta accttgatga
     2041 agaggtgaat gtaattcctc cacacactcc agttaggact gttatgaaca ctatccaaca
     2101 attaatgatg attttaaatt cagcaagtga tcaaccttca gaaaatctga tttcctattt
     2161 taacaactgc acagtgaatc caaaagaaag tatactgaaa agagtgaagg atataggata
     2221 catctttaaa gagaaatttg ctaaagctgt gggacagggt tgtgtcgaaa ttggatcaca
     2281 gcgatacaaa cttggagttc gcttgtatta ccgagtaatg gaatccatgc ttaaatcaga
     2341 agaagaacga ttatccattc aaaattttag caaacttctg aatgacaaca tttttcatat
     2401 gtctttattg gcgtgcgctc ttgaggttgt aatggccaca tatagcagaa gtacatctca
     2461 gaatcttgat tctggaacag atttgtcttt cccatggatt ctgaatgtgc ttaatttaaa
     2521 agcctttgat ttttacaaag tgatcgaaag ttttatcaaa gcagaaggca acttgacaag
     2581 agaaatgata aaacatttag aacgatgtga acatcgaatc atggaatccc ttgcatggct
     2641 ctcagattca cctttatttg atcttattaa acaatcaaag gaccgagaag gaccaactga
     2701 tcaccttgaa tctgcttgtc ctcttaatct tcctctccag aataatcaca ctgcagcaga
     2761 tatgtatctt tctcctgtaa gatctccaaa gaaaaaaggt tcaactacgc gtgtaaattc
     2821 tactgcaaat gcagagacac aagcaacctc agccttccag acccagaagc cattgaaatc
     2881 tacctctctt tcactgtttt ataaaaaagt gtatcggcta gcctatctcc ggctaaatac
     2941 actttgtgaa cgccttctgt ctgagcaccc agaattagaa catatcatct ggaccctttt
     3001 ccagcacacc ctgcagaatg agtatgaact catgagagac aggcatttgg accaaattat
     3061 gatgtgttcc atgtatggca tatgcaaagt gaagaatata gaccttaaat tcaaaatcat
     3121 tgtaacagca tacaaggatc ttcctcatgc tgttcaggag acattcaaac gtgttttgat
     3181 caaagaagag gagtatgatt ctattatagt attctataac tcggtcttca tgcagagact
     3241 gaaaacaaat attttgcagt atgcttccac caggccccct accttgtcac caatacctca
     3301 cattcctcga agcccttaca agtttcctag ttcaccctta cggattcctg gagggaacat
     3361 ctatatttca cccctgaaga gtccatataa aatttcagaa ggtctgccaa caccaacaaa
     3421 aatgactcca agatcaagaa tcttagtatc aattggtgaa tcattcggga cttctgagaa
     3481 gttccagaaa ataaatcaga tggtatgtaa cagcgaccgt gtgctcaaaa gaagtgctga
     3541 aggaagcaac cctcctaaac cactgaaaaa actacgcttt gatattgaag gatcagatga
     3601 agcagatgga agtaaacatc tcccaggaga gtccaaattt cagcagaaac tggcagaaat
     3661 gacttctact cgaacacgaa tgcaaaagca gaaaatgaat gatagcatgg atacctcaaa
     3721 caaggaagag aaatgaggta ccgcggccgc ggggatccag acatgataag atacattgat
     3781 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
     3841 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     3901 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggttttttc ggatcctcta
     3961 gagtcgatct gcaggcatgc tagcttggcg taatcatggt catagctgtt tcctgtgtga
     4021 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     4081 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     4141 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     4201 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     4261 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     4321 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     4381 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     4441 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     4501 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     4561 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     4621 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4681 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4741 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4801 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     4861 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4921 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4981 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     5041 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     5101 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     5161 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     5221 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     5281 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     5341 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     5401 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     5461 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     5521 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     5581 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     5641 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     5701 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     5761 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     5821 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     5881 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     5941 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     6001 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6061 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6121 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca
     6181 ttaacctata aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac
     6241 ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat
     6301 gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg
     6361 cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata
     6421 ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt caggctgcgc
     6481 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg
     6541 ggatgtgctg caag
//
