LOCUS       sequence-484475-        4981 bp    DNA     circular SYN 25-AUG-2025
DEFINITION  Gateway middle entry clone containing mCerulean3-TUBB1A; Cyan 
            fluorescent reporter for visualizing tubulin in cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4981)
  AUTHORS   Gillespie W, Zhang Y, Ruiz OE, Cerda J, Ortiz-Guzman J, Turner WD, 
            Largoza G, Sherman M, 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 with diverse applications in vertebrate model 
            systems.
  JOURNAL   bioRxiv [Preprint]. 2024 Jul 13:2024.07.13.603267. doi: 
            10.1101/2024.07.13.603267.
  PUBMED    39026881
REFERENCE   2  (bases 1 to 4981)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 25, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2024 Jul 13:2024.07.13.603267. doi: 
            10.1101/2024.07.13.603267."
COMMENT     Sequence Label: pME-mCerulean3-TUBB1A (JDW 1508)
FEATURES             Location/Qualifiers
     source          1..4981
                     /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"
     regulatory      696..705
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(702..704,705..707,708..1418)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(747..768)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1008..1027)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1355..1376
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2958..3079
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2995..3014)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3049..3068
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(3106..3205)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3222..3241)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3223..3241)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3246..3262)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3246..3262)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3364..3383)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3375..4184
                     /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(3442..3461)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4331..4919
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4820..4839
                     /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 agcaggctgc aggtacctta attaactcga ggcacgccac catggtgagc aagggcgagg
      721 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
      781 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
      841 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgagct
      901 ggggcgtgca gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
      961 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     1021 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     1081 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaacgcca
     1141 tccacggcaa cgtctatatc accgccgaca agcagaagaa cggcatcaag gccaacttcg
     1201 gcctcaactg caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     1261 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtcca
     1321 agctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     1381 ccgccgggat cactctcggc atggacgagc tgtacaagtc cggactcaga tctggcagcg
     1441 gtggaggcag cgcatccggc ggaagcggaa gcgtgcgtga gtgcatctcc atccacgttg
     1501 gccaggctgg tgtccagatt ggcaatgcct gctgggagct ctactgcctg gaacacggca
     1561 tccagcccga tggccagatg ccaagtgaca agaccattgg gggaggagat gactccttca
     1621 acaccttctt cagtgagacg ggcgctggca agcacgtgcc ccgggctgtg tttgtagact
     1681 tggaacccac agtcattgat gaagttcgca ctggcaccta ccgccagctc ttccaccctg
     1741 agcagctcat cacaggcaag gaagatgctg ccaataacta tgcccgaggg cactacacca
     1801 ttggcaagga gatcattgac cttgtgttgg accgaattcg caagctggct gaccagtgca
     1861 ccggtcttca gggcttcttg gttttccaca gctttggtgg gggaactggt tctgggttca
     1921 cctccctgct catggaacgt ctctcagttg attatggcaa gaagtccaag ctggagttct
     1981 ccatttaccc agcaccccag gtttccacag ctgtagttga gccctacaac tccatcctca
     2041 ccacccacac caccctggag cactctgatt gtgccttcat ggtagacaat gaggccatct
     2101 atgacatctg tcgtagaaac ctcgatatcg agcgcccaac ctacactaac cttaaccgcc
     2161 ttattagcca gattgtgtcc tccatcactg cttccctgag atttgatgga gccctgaatg
     2221 ttgacctgac agaattccag accaacctgg tgccctaccc ccgcatccac ttccctctgg
     2281 ccacatatgc ccctgtcatc tctgctgaga aagcctacca tgaacagctt tctgtagcag
     2341 agatcaccaa tgcttgcttt gagccagcca accagatggt gaaatgtgac cctcgccatg
     2401 gtaaatacat ggcttgctgc ctgttgtacc gtggtgacgt ggttcccaaa gatgtcaatg
     2461 ctgccattgc caccatcaaa accaagcgca gcatccagtt tgtggattgg tgccccactg
     2521 gcttcaaggt tggcatcaac taccagcctc ccactgtggt gcctggtgga gacctggcca
     2581 aggtacagag agctgtgtgc atgctgagca acaccacagc cattgctgag gcctgggctc
     2641 gcctggacca caagtttgac ctgatgtatg ccaagcgtgc ctttgttcac tggtacgtgg
     2701 gtgaggggat ggaggaaggc gagttttcag aggcccgtga agatatggct gcccttgaga
     2761 aggattatga ggaggttggt gtggattctg ttgaaggaga gggtgaggaa gaaggagagg
     2821 aatactaagg atccaccgga tctagataac tgatcataat cagccatacc acatttgtag
     2881 aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga
     2941 atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     3001 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     3061 aactcatcaa tgtatcttaa cgactagtgc ggccgcaagc tttgcaccca gctttcttgt
     3121 acaaagttgg cattataaga aagcattgct tatcaatttg ttgcaacgaa caggtcacta
     3181 tcagtcaaaa taaaatcatt atttgccatc cagctgatat cccctatagt gagtcgtatt
     3241 acatggtcat agctgtttcc tggcagctct ggcccgtgtc tcaaaatctc tgatgttaca
     3301 ttgcacaaga taaaaatata tcatcatgaa caataaaact gtctgcttac ataaacagta
     3361 atacaagggg tgttatgagc catattcaac gggaaacgtc gaggccgcga ttaaattcca
     3421 acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg
     3481 cgacaatcta tcgcttgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca
     3541 aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat
     3601 ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca
     3661 ccactgcgat ccccggaaaa acagcattcc aggtattaga agaatatcct gattcaggtg
     3721 aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta
     3781 attgtccttt taacagcgat cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata
     3841 acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag
     3901 tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg
     3961 atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg
     4021 gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg
     4081 agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata
     4141 tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaatcagaa ttggttaatt
     4201 ggttgtaaca ctggcagagc attacgctga cttgacggga cggcgcaagc tcatgaccaa
     4261 aatcccttaa cgtgagttac gcgtcgttcc actgagcgtc agaccccgta gaaaagatca
     4321 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     4381 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     4441 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     4501 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     4561 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     4621 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     4681 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     4741 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     4801 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     4861 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     4921 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     4981 t
//
