LOCUS       sequence-418734-        7428 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 7428)
  AUTHORS   van den Heuvel S, Harlow E
  TITLE     Distinct roles for cyclin-dependent kinases in cell cycle control.
  JOURNAL   Science 1993 Dec 24;262(5142):2050-4.
  PUBMED    8266103
REFERENCE   2  (bases 1 to 7428)
  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: "Science"; 
            date: "1993-12-24"; volume: "262(5142)"; pages: "2050-4"
COMMENT     Sequence Label: Cdk4
FEATURES             Location/Qualifiers
     source          1..7428
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(209..228)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    253..300
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     enhancer        593..972
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        973..1176
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1127..1147
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1173..1197
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          1288..1860
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     primer_bind     complement(2942..2961)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3007..3062
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3061..3080)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3420..3436)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3420..3436)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3433..3455)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3444..3460
                     /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(3468..3474,3475..3492,3493..3498))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3513..3534
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3651..3668)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3822..4410)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3902..3921)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4581..5372,5373..5441))
                     /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     5204..5223
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5442..5546)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5614..5632
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5670..5692)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5792..5811
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6005..6027
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6019..6036
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6020..6036
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        6447..6592
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     CDS             join(6649..7428,1..15)
                     /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(6703..6722)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7313..7332
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
       61 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
      121 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
      181 cccaccccgg gagatggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa
      241 cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc
      301 ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt
      361 ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag
      421 gcccagggct cgcagccaac gtcggggcgg caagccctgc catagccacg ggccccgtgg
      481 gttagggacg gcggatcgcg gccctctaga gagcttggcc cattgcatac gttgtatcca
      541 tatcataata tgtacattta tattggctca tgtccaacat taccgccatg ttgacattga
      601 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
      661 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
      721 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      781 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      841 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      901 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      961 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
     1021 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     1081 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
     1141 aggcgtgtac ggtgggaggt ctatataagc agagctcgtt tagtgaaccg tcagatcgcc
     1201 tggagacgcc atccacgctg ttttgacctc catagaagac accgggaccg atccagcctc
     1261 cggtcgaccg atcctgagaa cttcagggtg agtttgggga cccttgattg ttctttcttt
     1321 ttcgctattg taaaattcat gttatatgga gggggcaaag ttttcagggt gttgtttaga
     1381 atgggaagat gtcccttgta tcaccatgga ccctcatgat aattttgttt ctttcacttt
     1441 ctactctgtt gacaaccatt gtctcctctt attttctttt cattttctgt aactttttcg
     1501 ttaaacttta gcttgcattt gtaacgaatt tttaaattca cttttgttta tttgtcagat
     1561 tgtaagtact ttctctaatc actttttttt caaggcaatc agggtatatt atattgtact
     1621 tcagcacagt tttagagaac aattgttata attaaatgat aaggtagaat atttctgcat
     1681 ataaattctg gctggcgtgg aaatattctt attggtagaa acaactacac cctggtcatc
     1741 atcctgcctt tctctttatg gttacaatga tatacactgt ttgagatgag gataaaatac
     1801 tctgagtcca aaccgggccc ctctgctaac catgttcatg ccttcttctc tttcctacag
     1861 ctcctgggca acgtgctggt tgttgtgctg tctcatcatt ttggcaaaga attcctcgac
     1921 ggatccacca tggctacctc tcgatatgag ccagtggctg aaattggtgt cggtgcctat
     1981 gggacagtgt acaaggcccg tgatccccac agtggccact ttgtggccct caagagtgtg
     2041 agagtcccca atggaggagg aggtggagga ggccttccca tcagcacagt tcgtgaggtg
     2101 gctttactga ggcgactgga ggcttttgag catcccaatg ttgtccggct gatggacgtc
     2161 tgtgccacat cccgaactga ccgggagatc aaggtaaccc tggtgtttga gcatgtagac
     2221 caggacctaa ggacatatct ggacaaggca cccccaccag gcttgccagc cgaaacgatc
     2281 aaggatctga tgcgccagtt tctaagaggc ctagatttcc ttcatgccaa ttgcatcgtt
     2341 caccgagatc tgaagccaga gaacattctg gtgacaagtg gtggaacagt caagctggct
     2401 gactttggcc tggccagaat ctacagctac cagatggcac ttacacccgt ggttgttaca
     2461 ctctggtacc gagctcccga agttcttctg cagtccacat atgcaacacc tgtggacatg
     2521 tggagtgttg gctgtatctt tgcagagatg tttcgtcgaa agcctctctt ctgtggaaac
     2581 tctgaagccg accagttggg caaaatcttt gacctgattg ggctgcctcc agaggatgac
     2641 tggcctcgag atgtatccct gccccgtgga gcctttcccc ccagagggcc ccgcccagtg
     2701 cagtcggtgg tacctgagat ggaggagtcg ggagcacagc tgctgctgga aatgctgact
     2761 tttaacccac acaagcgaat ctctgccttt cgagctctgc agcactctta tctacataag
     2821 gatgaaggta atccggagtg agcaatggat ccgtcgagga attcactcct caggtgcagg
     2881 ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct ggctcacaaa taccactgag
     2941 atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag catctgactt
     3001 ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct
     3061 cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt
     3121 agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg ctataaagag
     3181 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga
     3241 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa
     3301 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca
     3361 tagctgtccc tcttctctta tggagatccc tcgacggatc ggccgcaatt cgtaatcatg
     3421 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     3481 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     3541 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     3601 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     3661 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     3721 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     3781 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     3841 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     3901 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     3961 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     4021 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     4081 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     4141 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     4201 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     4261 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     4321 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     4381 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     4441 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     4501 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     4561 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     4621 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     4681 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     4741 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     4801 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     4861 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     4921 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     4981 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     5041 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     5101 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     5161 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     5221 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     5281 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     5341 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     5401 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     5461 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     5521 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     5581 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt
     5641 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
     5701 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     5761 gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg
     5821 caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc
     5881 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
     5941 ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg
     6001 ttttcccagt cacgacgttg taaaacgacg gccagtgcca agcttcacgc tgccgcaagc
     6061 actcagggcg caagggctgc taaaggaagc ggaacacgta gaaagccagt ccgcagaaac
     6121 ggtgctgacc ccggatgaat gtcagctact gggctatctg gacaagggaa aacgcaagcg
     6181 caaagagaaa gcaggtagct tgcagtgggc ttacatggcg atagctagac tgggcggttt
     6241 tatggacagc aagcgaaccg gaattgccag ctggggcgcc ctctggtaag gttgggaagc
     6301 cctgcaaagt aaactggatg gctttcttgc cgccaaggat ctgatggcgc aggggatcaa
     6361 gatcctgctt catccccgtg gcccgttgct cgcgtttgct ggcggtgtcc ccggaagaaa
     6421 tatatttgca tgtctttagt tctatgatga cacaaacccc gcccagcgtc ttgtcattgg
     6481 cgaattcgaa cacgcagatg cagtcggggc ggcgcggtcc caggtccact tcgcatatta
     6541 aggtgacgcg tgtggcctcg aacaccgagc gaccctgcag cgacccgctt aacagcgtca
     6601 acagcgtgcc gcagatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     6661 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     6721 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     6781 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     6841 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     6901 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     6961 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     7021 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     7081 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     7141 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     7201 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     7261 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     7321 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     7381 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttctt
//
