LOCUS       sequence-446725-        3699 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A Gateway compatible middle entry clone containing a histone H2B 
            fusion to mCerulean to label the nucleus .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3699)
  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 3699)
  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: 
            "Development."; date: "2025-11-15"; volume: "152(22):dev204308. 
            doi"; pages: " 10.1242/dev.204308. Epub 2025 Nov 13"
COMMENT     Sequence Label: pME H2B mCerulean (JDW 1150)
FEATURES             Location/Qualifiers
     source          1..3699
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             692..1069
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     regulatory      1082..1091
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1088..1090,1091..1093,1094..1807)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1133..1154)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1394..1413)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1741..1762
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1824..1923)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1940..1959)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1941..1959)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1964..1980)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1964..1980)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2082..2101)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2093..2902
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2160..2179)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3049..3637
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3538..3557
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctct agagaattcg gtaccgccac catgccagag ccagcgaagt ctgctcccgc
      721 cccgaaaaag ggctccaaga aggcggtgac taaggcgcag aagaaaggcg gcaagaagcg
      781 caagcgcagc cgcaaggaga gctattccat ctatgtgtac aaggttctga agcaggtcca
      841 ccctgacacc ggcatttcgt ccaaggccat gggcatcatg aattcgtttg tgaacgacat
      901 tttcgagcgc atcgcaggtg aggcttcccg cctggcgcat tacaacaagc gctcgaccat
      961 cacctccagg gagatccaga cggccgtgcg cctgctgctg cctggggagt tggccaagca
     1021 cgccgtgtcc gagggtacta aggccatcac caagtacacc agcgctaagg atccaccggt
     1081 cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     1141 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     1201 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     1261 gcccaccctc gtgaccaccc tgacctgggg cgtgcagtgc ttcgcccgct accccgacca
     1321 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     1381 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     1441 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     1501 ggggcacaag ctggagtaca acgccatcag cgacaacgtc tatatcaccg ccgacaagca
     1561 gaagaacggc atcaaggcca acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     1621 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     1681 caaccactac ctgagcaccc agtccaagct gagcaaagac cccaacgaga agcgcgatca
     1741 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     1801 caagtaataa gcggccgcct cgagcccagc tttcttgtac aaagttggca ttataagaaa
     1861 gcattgctta tcaatttgtt gcaacgaaca ggtcactatc agtcaaaata aaatcattat
     1921 ttgccatcca gctgatatcc cctatagtga gtcgtattac atggtcatag ctgtttcctg
     1981 gcagctctgg cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc
     2041 atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca
     2101 tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg
     2161 gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg
     2221 gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt
     2281 tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa
     2341 gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac
     2401 agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc
     2461 agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg
     2521 cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga
     2581 ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact
     2641 tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat
     2701 ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg
     2761 ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa
     2821 acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt
     2881 gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacact ggcagagcat
     2941 tacgctgact tgacgggacg gcgcaagctc atgaccaaaa tcccttaacg tgagttacgc
     3001 gtcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     3061 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     3121 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     3181 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     3241 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     3301 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     3361 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     3421 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     3481 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     3541 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     3601 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     3661 acggttcctg gccttttgct ggccttttgc tcacatgtt
//
