LOCUS       sequence-252608-        6554 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of human Rb with all Ser/Thr Cdk acceptor sites
            removed.
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 delta CDK
FEATURES             Location/Qualifiers
     source          1..6554
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(126..714)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(206..225)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(885..1676,1677..1745))
                     /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     1508..1527
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1746..1850)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1918..1936
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1974..1996)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2096..2115
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2309..2331
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2323..2340
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2324..2340
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        2367..2670
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2671..2874
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2825..2845
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2871..2895
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          3012..3108
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             3172..3198
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    6033..6167
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6057..6076)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6111..6130
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(6278..6294)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6278..6294)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6291..6313)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6302..6318
                     /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(6326..6332,6333..6350,6351..6356))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6371..6392
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6509..6526)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
       61 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
      121 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
      181 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
      241 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
      301 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
      361 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
      421 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
      481 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
      541 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
      601 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
      661 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
      721 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
      781 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
      841 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
      901 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
      961 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     1021 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     1081 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     1141 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     1201 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     1261 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     1321 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     1381 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     1441 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     1501 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     1561 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     1621 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     1681 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     1741 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     1801 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     1861 cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat
     1921 aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga
     1981 cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa
     2041 gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca
     2101 tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta
     2161 aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg
     2221 cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg
     2281 cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt
     2341 gagttcgagc ttgcatgcct gcaggtcgtt acataactta cggtaaatgg cccgcctggc
     2401 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     2461 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     2521 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     2581 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     2641 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     2701 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     2761 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     2821 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta
     2881 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac
     2941 cgggaccgat ccagcctccg gactctagag gatccggtac tagaggaact gaaaaaccag
     3001 aaagttaact ggtaagttta gtctttttgt cttttatttc aggtcccgga tccggtggtg
     3061 gtgcaaatca aagaactgct cctcagtgga tgttgccttt acttctaggc ctgtacggaa
     3121 gtgttacttc tgctctaaaa gctgcggaat tgtacccgcg ggcccaccat gtacccatac
     3181 gatgttccag attacgctct tatggccatg gaggcccgaa ttcggtcgac cccgcccaaa
     3241 gccccccgaa aaacggccgc caccgccgcc gctgccgccg cggaaccccc ggcaccgccg
     3301 ccgccgcccc ctcctgagga ggacccagag caggacagcg gcccggagga cctgcctctc
     3361 gtcaggcttg agtttgaaga aacagaagaa cctgatttta ctgcattatg tcagaaatta
     3421 aagataccag atcatgtcag agagagagct tggttaactt gggagaaagt ttcatctgtg
     3481 gatggagtat tgggaggtta tattcaaaag aaaaaggaac tgtggggaat ctgtatcttt
     3541 attgcagcag ttgacctaga tgagatgtcg ttcactttta ctgagctaca gaaaaacata
     3601 gaaatcagtg tccataaatt ctttaactta ctaaaagaaa ttgataccag taccaaagtt
     3661 gataatgcta tgtcaagact gttgaagaag tatgatgtat tgtttgcact cttcagcaaa
     3721 ttggaaagga catgtgaact tatatatttg acacaaccca gcagttcgat atctactgaa
     3781 ataaattctg cattggtgct aaaagtttct tggatcacat ttttattagc taaaggggaa
     3841 gtattacaaa tggaagatga tctggtgatt tcatttcagt taatgctatg tgtccttgac
     3901 tattttatta aactcgcacc tcccatgttg ctcaaagaac catataaaac agctgttata
     3961 cccattaatg gtgcacctcg agcacccagg cgaggtcaga acaggagtgc acggatagca
     4021 aaacaactag aaaatgatac aagaattatt gaagttctct gtaaagaaca tgaatgtaat
     4081 atagatgagg tgaaaaatgt ttatttcaaa aattttatac cttttatgaa ttctcttgga
     4141 cttgtaacat ctaatggact tccagaggtt gaaaatcttt ctaaacgata cgaagaaatt
     4201 tatcttaaaa ataaagatct agatgcaaga ttatttttgg atcatgataa aactcttcag
     4261 actgattcta tagacagttt tgaaacacag agagcaccac gaaaaagtaa ccttgatgaa
     4321 gaggtgaatg taattcctcc acacgctcca gttaggactg ttatgaacac tatccaacaa
     4381 ttaatgatga ttttaaattc agcaagtgat caaccttcag aaaatctgat ttcctatttt
     4441 aacaactgca cagtgaatcc aaaagaaagt atactgaaaa gagtgaagga tataggatac
     4501 atctttaaag agaaatttgc taaagctgtg ggacagggtt gtgtcgaaat tggatcacag
     4561 cgatacaaac ttggagttcg cttgtattac cgagtaatgg aatccatgct taaatcagaa
     4621 gaagaacgat tatccattca aaattttagc aaacttctga atgacaacat ttttcatatg
     4681 tctttattgg cgtgcgctct tgaggttgta atggccacat atagcagaag tacatctcag
     4741 aatcttgatt ctggaacaga tttgtctttc ccatggattc tgaatgtgct taatttaaaa
     4801 gcctttgatt tttacaaagt gatcgaaagt tttatcaaag cagaaggcaa cttgacaaga
     4861 gaaatgataa aacatttaga acgatgtgaa catcgaatca tggaatccct tgcatggctc
     4921 tcagattcac ctttatttga tcttattaaa caatcaaagg accgagaagg accaactgat
     4981 caccttgaat ctgcttgtcc tcttaatctt cctctccaga ataatcacac tgcagcagat
     5041 atgtatcttg ctcctgtaag agctccaaag aaaaaaggtt caactacgcg tgtaaattct
     5101 actgcaaatg cagagacaca agcaacctca gccttccaga cccagaagcc attgaaatct
     5161 acctctcttt cactgtttta taaaaaagtg tatcggctag cctatctccg gctaaataca
     5221 ctttgtgaac gccttctgtc tgagcaccca gaattagaac atatcatctg gacccttttc
     5281 cagcacaccc tgcagaatga gtatgaactc atgagagaca ggcatttgga ccaaattatg
     5341 atgtgttcca tgtatggcat atgcaaagtg aagaatatag accttaaatt caaaatcatt
     5401 gtaacagcat acaaggatct tcctcatgct gttcaggaga cattcaaacg tgttttgatc
     5461 aaagaagagg agtatgattc tattatagta ttctataact cggtcttcat gcagagactg
     5521 aaaacaaata ttttgcagta tgcttccacc aggcccccta ccttggcacc aatacctcac
     5581 attcctcgag ccccttacaa gtttcctagt gcacccttac ggattcctgg agggaacatc
     5641 tatattgcac ccctgaaggc tccatataaa atttcagaag gtctgccagc accaacaaaa
     5701 atggctccaa gatcaagaat cttagtatca attggtgaat cattcgggac ttctgagaag
     5761 ttccagaaaa taaatcagat ggtatgtaac agcgaccgtg tgctcaaaag aagtgctgaa
     5821 ggaagcaacc ctcctaaacc actgaaaaaa ctacgctttg atattgaagg atcagatgaa
     5881 gcagatggaa gtaaacatct cccaggagag tccaaatttc agcagaaact ggcagaaatg
     5941 acttctactc gaacacgaat gcaaaagcag aaaatgaatg atagcatgga tacctcaaac
     6001 aaggaagaga aatgaggtac cgcggccgcg gggatccaga catgataaga tacattgatg
     6061 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     6121 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     6181 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttcg gatcctctag
     6241 agtcgatctg caggcatgct agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     6301 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     6361 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     6421 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     6481 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     6541 ggctgcggcg agcg
//
