LOCUS       sequence-495403-        7017 bp    DNA     circular SYN 16-DEC-2025
DEFINITION  A CAGGS driven expression vector containing FastFUCCI to label cell 
            cycle state. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7017)
  AUTHORS   Gillespie WB 3rd, Zhang Y, Ruiz OE, Cerda J 3rd, Ortiz-Guzman J, 
            Sherman M, Turner WD, Largoza G, Mosser LE, Fujimoto E, Chien CB, 
            Kwan KM, Arenkiel BR, Devine WP, Wythe JD
  TITLE     MultiSite Assembly of Gateway Induced Clones (MAGIC): a flexible 
            cloning toolbox for use in vertebrate model systems.
  JOURNAL   Development. 2025 Nov 15;152(22):dev204308. doi: 10.1242/dev.204308.
            Epub 2025 Nov 13.
  PUBMED    41230856
REFERENCE   2  (bases 1 to 7017)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 16, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Development."; date: "2025-11-15"; volume: "152(22):dev204308. 
            doi"; pages: " 10.1242/dev.204308. Epub 2025 Nov 13"
COMMENT     Sequence Label: pCAGEN-FastFUCCI (JDW 1494)
FEATURES             Location/Qualifiers
     source          1..7017
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..662
                     /label=chicken beta-actin promoter
     intron          663..1679
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1603..1625
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1687..1706
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1746..1770
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1812..2462
                     /codon_start=1
                     /product="monomeric Kusabira-Orange 2 fluorescent protein"
                     /label=mKO2
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAH"
     CDS             2517..2789
                     /codon_start=1
                     /product="degron consisting of residues 30-120 of human 
                     Cdt1 (Zielke and Edgar, 2015)"
                     /label=Cdt1 (30-120)
                     /translation="PSPARPALRAPASATSGSRKRARPPAAPGRDQARPPARRRLRLSV
                     DEVSSPSTPEAPDIPACPSPGQKIKKSTPAAGQPPHLTSAQDQDTI"
     CDS             2796..2849
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2856..3533
                     /codon_start=1
                     /product="humanized monomeric Azami-Green fluorescent 
                     protein"
                     /label=hmAzami-Green
                     /note="mammalian codon-optimized"
                     /translation="MVSVIKPEMKIKLCMRGTVNGHNFVIEGEGKGNPYEGTQILDLNV
                     TEGAPLPFAYDILTTVFQYGNRAFTKYPADIQDYFKQTFPEGYHWERSMTYEDQGICTA
                     TSNISMRGDCFFYDIRFDGTNFPPNGPVMQKKTLKWEPSTEKMYVEDGVLKGDVNMRLL
                     LEGGGHYRCDFKTTYKAKKEVRLPDAHKIDHRIEILKHDKDYNKVKLYENAVARYSMLP
                     SQAK"
     CDS             2856..3533
                     /codon_start=1
                     /product="monomeric Azami-Green fluorescent protein 
                     (Karasawa et al., 2003)"
                     /label=mAzami-Green
                     /translation="MVSVIKPEMKIKLCMRGTVNGHNFVIEGEGKGNPYEGTQILDLNV
                     TEGAPLPFAYDILTTVFQYGNRAFTKYPADIQDYFKQTFPEGYHWERSMTYEDQGICTA
                     TSNISMRGDCFFYDIRFDGTNFPPNGPVMQKKTLKWEPSTEKMYVEDGVLKGDVNMRLL
                     LEGGGHYRCDFKTTYKAKKEVRLPDAHKIDHRIEILKHDKDYNKVKLYENAVARYSMLP
                     SQAK"
     CDS             3564..3893
                     /codon_start=1
                     /product="degron consisting of residues 1-110 of human 
                     Geminin (Zielke and Edgar, 2015)"
                     /label=Gem1 (1-110)
                     /translation="MNPSMKQKQEEIKENIKNSSVPRRTLKMIQPSASGSLVGRENELS
                     AGLSKRKHRNDHLTSTTSSPGVIVPESSENKNLGGVTQESFDLMIKENPSSQYWKEVAE
                     KRRKAL"
     protein_bind    complement(3923..3947)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(4039..4058)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4104..4159
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4158..4177)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4520..4536)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4520..4536)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4533..4555)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4544..4560
                     /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(4568..4574,4575..4592,4593..4598))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4613..4634
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4693..4888
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4739..4874
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4894..5028
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4931..4950)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4985..5004
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5096..5113)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5267..5855)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5347..5366)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6026..6817,6818..6886))
                     /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     6649..6668
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6887..6991)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg ggggggcgcg cgccgggggg gggggggcgg ggcgaggggc
      541 ggggcggggc gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt
      601 ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg
      661 cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc
      721 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
      781 ctccgggctg taattagcgc ttggtttaat gacggcttgt ttcttttctg tggctgcgtg
      841 aaagccttga ggggctccgg gagggccctt tgtgcggggg gagcggctcg gggggtgcgt
      901 gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc ggctgtgagc
      961 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt gcgcgagggg agcgcggccg
     1021 ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa caaaggctgc gtgcggggtg
     1081 tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt cgggctgcaa ccccccctgc
     1141 acccccctcc ccgagttgct gagcacggcc cggcttcggg tgcggggctc cgtacggggc
     1201 gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca ggtgggggtg ccgggcgggg
     1261 cggggccgcc tcgggccggg gagggctcgg gggaggggcg cggcggcccc cggagcgccg
     1321 gcggctgtcg aggcgcggcg agccgcagcc attgcctttt atggtaatcg tgcgagaggg
     1381 cgcagggact tcctttgtcc caaatctgtg cggagccgaa atctgggagg cgccgccgca
     1441 ccccctctag cgggcgcggg gcgaagcggt gcggcgccgg caggaaggaa atgggcgggg
     1501 agggccttcg tgcgtcgccg cgccgccgtc cccttctccc tctccagcct cggggctgtc
     1561 cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg
     1621 tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     1681 tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttcctcgagg
     1741 atatcacaag tttgtacaaa aaagcaggct ggtctgactg gtacctcgtg attaattaag
     1801 aattgggatc catggtgtcc gtgatcaagc ccgagatgaa gatgcggtac tacatggacg
     1861 gcagcgtgaa cggccacgag ttcaccatcg agggcgaggg caccggcaga ccttacgagg
     1921 gccaccagga aatgaccctg agagtcacca tggccgaggg cggacccatg cccttcgcct
     1981 tcgacctggt gtcccacgtg ttctgctacg gccaccgggt gttcaccaag taccccgagg
     2041 aaatccccga ctacttcaag caggccttcc ccgagggcct gagctgggag cggagcctgg
     2101 aattcgagga tggcggcagc gcctctgtgt ctgcccacat cagcctgcgg ggcaacacct
     2161 tctaccacaa gagcaagttc accggcgtga acttccccgc cgacggcccc atcatgcaga
     2221 accagagcgt ggactgggag cccagcaccg agaagatcac cgccagcgac ggcgtgctga
     2281 agggcgacgt gaccatgtac ctgaagctgg aaggcggcgg aaaccacaag tgccagatga
     2341 aaaccaccta caaggccgcc aaagaaatcc tggaaatgcc cggcgaccac tacatcggcc
     2401 accggctggt gcggaaaacc gagggcaaca tcaccgagca ggtggaggat gccgtggccc
     2461 actacagcat gctgcctagc ctcggcggct accctagcca ttggaggccc ctggaaccct
     2521 ctcctgccag acctgccctg agagcccctg cctccgccac ctccggcagc agaaagagag
     2581 ccagacctcc tgccgcccct ggcagagatc aggccaggcc ccctgcccgc agacggctcc
     2641 ggctgtccgt ggatgaggtg tccagcccta gcacccccga ggcccccgat atccctgcct
     2701 gtcccagccc tggccagaaa atcaagaaga gcacacctgc cgccggacag cctcctcacc
     2761 tgaccagcgc ccaggaccag gacaccatcg tcgacgaggg caggggcagc ctgctgacct
     2821 gcggcgacgt ggaggagaac cccggcccca gatccatggt gtccgtgatc aagcccgaga
     2881 tgaagatcaa gctgtgcatg cggggaaccg tgaacggcca caacttcgtg atcgagggcg
     2941 agggcaaggg aaacccctac gagggcaccc agatcctgga cctgaacgtg accgaaggcg
     3001 cccctctgcc cttcgcctac gacatcctga ccaccgtgtt ccagtacggc aaccgggcct
     3061 tcaccaagta ccccgccgac atccaggact acttcaagca gaccttcccc gagggctacc
     3121 actgggagcg gagcatgacc tacgaggacc agggcatctg caccgccacc tccaacatca
     3181 gcatgagggg cgactgcttt ttctacgaca tcagattcga cggcaccaac ttccccccca
     3241 acggccccgt gatgcagaaa aagaccctga agtgggagcc cagcaccgag aagatgtacg
     3301 tggaggacgg cgtgctgaag ggcgacgtga acatgcggct gctgctggaa ggcggcggac
     3361 actacagatg cgacttcaag accacctaca aggccaagaa agaagtgcgg ctgcccgacg
     3421 cccacaagat cgaccaccgg atcgagatcc tgaagcacga caaggactac aacaaagtga
     3481 agctgtacga gaacgccgtg gccaggtaca gcatgctgcc cagccaggcc aagggctacc
     3541 ccagccactg gcggcctctg gaaatgaacc ccagcatgaa gcagaagcag gaagagatca
     3601 aagagaacat caagaacagc agcgtgccca ggcggaccct gaagatgatc cagcccagcg
     3661 ccagcggatc tctggtgggc cgggagaatg agctgtctgc cggcctgagc aagcggaagc
     3721 accggaacga ccacctgacc agcaccacct ctagcccagg cgtgatcgtg cccgagagca
     3781 gcgagaacaa gaacctgggc ggcgtgaccc aagaaagctt cgacctgatg atcaaagaaa
     3841 accccagcag ccagtactgg aaagaggtgg ccgagaagcg gcggaaggcc ctgtagagat
     3901 ctgactgagc ggccgccagt caacccagct ttcttgtaca aagtggtgat atcgcggccg
     3961 cactcctcag gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca atgccctggc
     4021 tcacaaatac cactgagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc
     4081 ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg
     4141 gaattttttg tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca
     4201 gaatgagtat ttggtttaga gtttggcaac atatgcccat atgctggctg ccatgaacaa
     4261 aggttggcta taaagaggtc atcagtatat gaaacagccc cctgctgtcc attccttatt
     4321 ccatagaaaa gccttgactt gaggttagat tttttttata ttttgttttg tgttattttt
     4381 ttctttaaca tccctaaaat tttccttaca tgttttacta gccagatttt tcctcctctc
     4441 ctgactactc ccagtcatag ctgtccctct tctcttatgg agatccctcg acctgcagcc
     4501 caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     4561 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     4621 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4681 gccagcggat ccgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     4741 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     4801 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     4861 cttttttgga ggcctaggct tttgcaaaaa gctaacttgt ttattgcagc ttataatggt
     4921 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     4981 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggatccg ctgcattaat
     5041 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     5101 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     5161 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     5221 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     5281 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5341 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5401 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5461 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5521 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5581 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     5641 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     5701 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     5761 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5821 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     5881 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5941 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     6001 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     6061 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6121 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6181 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     6241 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6301 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6361 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6421 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6481 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6541 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6601 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     6661 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6721 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6781 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6841 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6901 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6961 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctg
//
