LOCUS       sequence-156475-        7893 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7893)
  AUTHORS   Hinds PW, Mittnacht S, Dulic V, Arnold A, Reed SI, Weinberg RA
  TITLE     Regulation of retinoblastoma protein functions by ectopic expression
            of human cyclins.
  JOURNAL   Cell. 1992 Sep 18. 70(6):993-1006.
  PUBMED    1388095
REFERENCE   2  (bases 1 to 7893)
  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: "Cell. 1992 
            Sep 18. 70(6):993-1006."
COMMENT     Sequence Label: Rc/CMV cyclin E
FEATURES             Location/Qualifiers
     source          1..7893
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(11..28)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1981..2000)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2172..2551
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2552..2755
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2706..2726
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2800..2819
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2800..2818
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(5289..5307)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5290..5307)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(5327..5344)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5333..5557
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5603..6116
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5690..5709)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5900..5921
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6139..6159)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6144..6473
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6324..6459
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6540..7334
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6594..6613)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7204..7223
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7508..7642
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7545..7564)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7599..7618
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7673..7689)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7673..7689)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7686..7708)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7697..7713
                     /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(7721..7727,7728..7745,7746..7751))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7766..7787
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag
     1981 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
     2041 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
     2101 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
     2161 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     2221 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     2281 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     2341 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     2401 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     2461 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     2521 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     2581 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     2641 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     2701 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctctg
     2761 gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc actataggga
     2821 gacccaagct tggtaccgag ctcggatcca ctagtaacgg ccgccagaat tccgggatgc
     2881 gaaggagcgg gacaccatga aggaggacgg cggcgcggag ttctcggctc gctccaggaa
     2941 gaggaaggca aacgtgaccg tttttttgca ggatccagat gaagaaatgg ccaaaatcga
     3001 caggacggcg agggaccagt gtgggagcca gccttgggac aataatgcag tctgtgcaga
     3061 cccctgctcc ctgatcccca cacctgacaa agaagatgat gaccgggttt acccaaactc
     3121 aacgtgcaag cctcggatta ttgcaccatc cagaggctcc ccgctgcctg tactgagctg
     3181 ggcaaataga gaggaagtct ggaaaatcat gttaaacaag gaaaagacat acttaaggga
     3241 tcagcacttt cttgagcaac accctcttct gcagccaaaa atgcgagcaa ttcttctgga
     3301 ttggttaatg gaggtgtgtg aagtctataa acttcacagg gagacctttt acttggcaca
     3361 agatttcttt gaccggtata tggcgacaca agaaaatgtt gtaaaaactc ttttacagct
     3421 tattgggatt tcatctttat ttattgcagc caaacttgag gaaatctatc ctccaaagtt
     3481 gcaccagttt gcgtatgtga cagatggagc ttgttcagga gatgaaattc tcaccatgga
     3541 attaatgatt atgaaggccc ttaagtggcg tttaagtccc ctgactattg tgtcctggct
     3601 gaatgtatac atgcaggttg catatctaaa tgacttacat gaagtgctac tgccgcagta
     3661 tccccagcaa atctttatac agattgcaga gctgttggat ctctgtgtcc tggatgttga
     3721 ctgccttgaa tttccttatg gtatacttgc tgcttcggcc ttgtatcatt tctcgtcatc
     3781 tgaattgatg caaaaggttt cagggtatca gtggtgcgac atagagaact gtgtcaagtg
     3841 gatggttcca tttgccatgg ttataaggga gacggggagc tcaaaactga agcacttcag
     3901 gggcgtcgct gatgaagatg cacacaacat acagacccac agagacagct tggatttgct
     3961 ggacaaagcc cgagcaaaga aagccatgtt gtctgaacaa aatagggctt ctcctctccc
     4021 cagtgggctc ctcaccccgc cacagagcgg taagaagcag agcagcgggc cggaaatggc
     4081 gtgaccaccc catccttctc caccaaagac agttgcgcgc ctgctccacg ttctcttctg
     4141 tctgttgcag cggaggcgtg cgtttgcttt tacagatatc tgaatggaag agtgtttctt
     4201 ccacaacaga agtatttctg tggatggcat caaacagggc aaagtgtttt ttattgaatg
     4261 cttataggtt ttttttaaat aagtgggtca agtacaccag ccacctccag acaccagtgc
     4321 gtgctcccga tgctgctatg gaaggtgcta cttgacctaa gggactccca caacaacaaa
     4381 agcttgaagc tgtggagggc cacggtggcg tggctctcct cgcaggtgtt ctgggctccg
     4441 ttgtaccaag tggagcaggt ggttgcgggc aagcgttgtg cagagcccat agccagctgg
     4501 gcagggggct gccctctcca cattatcagt tgacagtgta caatgccttt gatgaactgt
     4561 tttgtaagtg ctgctatatc tatccatttt ttaataaaga taatactgtt tttgaaaaaa
     4621 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaggaa gattatgtgg
     4681 tctaaagagc aaggctgcca gaagaactaa tccactatat atctcccata aacctacagt
     4741 cttgagtttg aagtctgcag caatgacttg agatttaagt aagtctttgg tgcccgtgaa
     4801 cagaatgcca aggacaaaga cataaacaaa acgagcttca ataataccct tggaaatgtc
     4861 cttgctgtcc ggccgccacg ggaaccggac actcccgatg gccgcccggt agcagtagac
     4921 gcccagcagc cccagcgcca gggcagcctt ggacacaggg gagcaccccc tctgcaccag
     4981 cacaaaaact acgaggaggg agagggcagc caggacagag agctcggctt tgtggtcaga
     5041 gctggtgagc cagtggccga ggtcaggccg gtgagcccac tgcacacctg tctggtttag
     5101 ggagcgcagc agccggcagc aggccagtat tagccacgga ctggccagca ccatccactt
     5161 ctcgcggcct ctgagcactt cggaggtaga ggggcttccg tggcctttgt ctcgctccag
     5221 gacatcacag ccaggcccgt cctgcggaat tctggcggcc gctcgagcat gcatctagag
     5281 ggccctattc tatagtgtca cctaaatgct agagctcgct gatcagcctc gactgtgcct
     5341 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     5401 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     5461 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     5521 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
     5581 tggggctcga ggggggatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     5641 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     5701 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     5761 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     5821 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     5881 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     5941 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     6001 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttaa
     6061 atatttgctt atacaatctt cctgtttttg gggcttttct gattatcaac cggggtgggt
     6121 accgagctcg aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     6181 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     6241 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     6301 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     6361 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     6421 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     6481 cgggagcttg gatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca
     6541 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     6601 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     6661 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     6721 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     6781 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     6841 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     6901 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     6961 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     7021 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg
     7081 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     7141 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     7201 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     7261 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     7321 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     7381 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     7441 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     7501 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     7561 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     7621 tgtatcttat catgtctgga tccgtcgacc tcgagagctt ggcgtaatca tggtcatagc
     7681 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     7741 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     7801 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     7861 gcgcggggag aggcggtttg cgtattgggc gct
//
