LOCUS       sequence-246298-        5135 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Contains insert allowing in-vivo visualization of CyclinB1 protein 
            (G2/M).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5135)
  AUTHORS   Tran M, Dave M, Lansford R
  TITLE     Fast and Efficient Expression of Multiple Proteins in Avian Embryos 
            Using mRNA Electroporation.
  JOURNAL   J Vis Exp. 2019 Jun 7;(148). doi: 10.3791/59664.
  PUBMED    31233018
REFERENCE   2  (bases 1 to 5135)
  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: "J Vis 
            Exp."; date: "2019-06-7"; volume: "(148). doi"; pages: " 
            10.3791/59664"
COMMENT     Sequence Label: pCS2+CyclinB1-GFP
FEATURES             Location/Qualifiers
     source          1..5135
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             join(427..429,430..432,433..1146)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(427..429,430..432,433..1143)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(472..493)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(733..752)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1080..1101
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1184..1318
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1208..1227)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1262..1281
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1431..1451)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1431..1449)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1470..1486)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1470..1486)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1483..1505)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1494..1510
                     /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(1518..1524,1525..1542,1543..1548))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1563..1584
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1701..1718)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1872..2460)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1952..1971)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2631..3422,3423..3491))
                     /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     3254..3273
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3492..3596)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3622..4077)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3759..3780)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3971..3990
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4151..5135
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atccaccatg gcgctcaggg tcactaggaa cacgaaaatt
      121 aacgcagaaa ataaggccaa ggtcagtatg gcaggcgcca agcgtgtgcc tgtgacagtt
      181 actgctgctt ccaagcccgg gctgagaccg agaactgctc ttggagacat tggtaataaa
      241 gtcagcgaag agctacaggc aagagtgcct ctgaaaaggg aagcaaaaac gctaggtact
      301 ggaaaaggta ctgttaaagc cctaccaaaa cctgtagaga aggtgcctgt gtgtgaacca
      361 gaggtggaac ttgctgagcc tgagcctgaa cctgaacttg aactagttca gaatttgtct
      421 agagccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
      481 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc
      541 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
      601 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
      661 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
      721 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
      781 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
      841 gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag
      901 aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
      961 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     1021 aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
     1081 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
     1141 aagtaagtcg agcctctaga actatagtga gtcgtattac gtagatccag acatgataag
     1201 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     1261 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     1321 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta
     1381 attcgcggcc gcggcgccaa tgcattgggc ccggtaccca gcttttgttc cctttagtga
     1441 gggttaattg cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     1501 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     1561 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     1621 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     1681 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     1741 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     1801 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     1861 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     1921 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     1981 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     2041 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     2101 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     2161 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     2221 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     2281 aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg
     2341 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     2401 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     2461 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     2521 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     2581 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     2641 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     2701 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     2761 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     2821 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     2881 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     2941 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     3001 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     3061 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     3121 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     3181 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     3241 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     3301 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     3361 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     3421 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     3481 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     3541 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     3601 tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
     3661 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
     3721 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
     3781 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
     3841 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
     3901 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
     3961 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
     4021 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc
     4081 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     4141 tacgccagtc gaccatagcc aattcaatat ggcgtatatg gactcatgcc aattcaatat
     4201 ggtggatctg gacctgtgcc aattcaatat ggcgtatatg gactcgtgcc aattcaatat
     4261 ggtggatctg gaccccagcc aattcaatat ggcggacttg gcaccatgcc aattcaatat
     4321 ggcggacttg gcactgtgcc aactggggag gggtctactt ggcacggtgc caagtttgag
     4381 gaggggtctt ggccctgtgc caagtccgcc atattgaatt ggcatggtgc caataatggc
     4441 ggccatattg gctatatgcc aggatcaata tataggcaat atccaatatg gccctatgcc
     4501 aatatggcta ttggccaggt tcaatactat gtattggccc tatgccatat agtattccat
     4561 atatgggttt tcctattgac gtagatagcc cctcccaatg ggcggtccca tataccatat
     4621 atggggcttc ctaataccgc ccatagccac tcccccattg acgtcaatgg tctctatata
     4681 tggtctttcc tattgacgtc atatgggcgg tcctattgac gtatatggcg cctcccccat
     4741 tgacgtcaat tacggtaaat ggcccgcctg gctcaatgcc cattgacgtc aataggacca
     4801 cccaccattg acgtcaatgg gatggctcat tgcccattca tatccgttct cacgccccct
     4861 attgacgtca atgacggtaa atggcccact tggcagtaca tcaatatcta ttaatagtaa
     4921 cttggcaagt acattactat tggaaggacg ccagggtaca ttggcagtac tcccattgac
     4981 gtcaatggcg gtaaatggcc cgcgatggct gccaagtaca tccccattga cgtcaatggg
     5041 gaggggcaat gacgcaaatg ggcgttccat tgacgtaaat gggcggtagg cgtgcctaat
     5101 gggaggtcta tataagcaat gctcgtttag ggaac
//
