LOCUS       sequence-447418-        7550 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Gateway compatible middle entry clone containing Sun1 reporter 
            fusion to 2 copies of sfGFP and 6 MYC tags (Nuclear envelope 
            reporter and affinity tag).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7550)
  AUTHORS   Gillespie W, Zhang Y, Ruiz OE, Cerda III J, Ortiz-Guzman J, Turner 
            WD, Largoza G, Sherman M, Mosser LE, Fujimoto E, Chien C-B, 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 2024.07.13.603267
REFERENCE   2  (bases 1 to 7550)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2024.07.13.603267"
COMMENT     Sequence Label: pME-Sun1-2xsfGFP-6xMYC-pA (JDW 681)
FEATURES             Location/Qualifiers
     source          1..7550
                     /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      750..759
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3576..3578,3579..3581,3582..4292)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4314..4316,4317..4319,4320..5030)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             5070..5099
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             5106..5135
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             5142..5171
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             5199..5228
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             5235..5264
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             5271..5300
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(5323..5340)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5329..5553
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(5675..5774)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(5791..5810)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5792..5810)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5815..5831)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5815..5831)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5933..5952)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5944..6753
                     /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(6011..6030)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      6900..7488
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7389..7408
                     /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 agcaggctac tagtggccgc tagcgtttgg gcttaagggg tctatcgatg ggtctgcggc
      721 cgcactactg gccggccact actacgcgtg ccaccatgga cttttctcgg ctgcacacgt
      781 acaccccacc ccagtgtgtg ccggagaaca ctggctacac ttacgcactc agttctagtt
      841 actcgtcgga tgctctggat tttgaaactg agcacaagtt ggaacctgta tttgactctc
      901 caaggatgtc ccgccgcagc ttgcgtctgg tcacaacagc ttcgtacagc agtggggaca
      961 gccaggctat tgattcgcac attagcacca gcagggccac ccccgccaag gggagagaaa
     1021 ccaggacagt caaacagaga agaagtgcaa gcaagccagc ttttagtatc aaccacctgt
     1081 cagggaaggg cttgtcctcg agcacaagcc atgacagctc ttgcagcctg cggagtgcca
     1141 cggtgctgcg gcaccctgtg ctagatgagt ccctgattcg tgagcagacc aaagtggacc
     1201 acttctgggg tctcgatgat gatggtgacc ttaaaggtgg aaataaagct gccactcagg
     1261 gaaatggtga actggcagca gaggtggcga gcagcaatgg atacacttgc cgtgactgca
     1321 ggatgctctc agcgcgcact gacgcactca cagcccactc tgccatccac gggaccacct
     1381 ccagggtgta ctccagagac aggactctca aaccacgcgg agtgtccttt tacctggata
     1441 ggactctgtg gctggccaag tccacctcct catcctttgc atcatttata gttcaacttt
     1501 tccaagtggt tttaatgaag ctcaattttg aaacttacaa attgaaaggc tatgaatcca
     1561 gagcttatga atcacagagc tatgagacaa agagccatga gtcagaagcc catctcggtc
     1621 actgtgggag gatgactgcc ggagaacttt ccagagtgga cggggagtcc ctgtgcgatg
     1681 actgtaaggg gaagaagcac cttgagatac acacagccac ccactcgcaa ctgccccagc
     1741 cacacagggt ggccggggcc atggggcgcc tctgcatcta tacaggtgac ctcttggttc
     1801 aagcactgcg aaggactaga gctgccgggt ggtctgtggc cgaggccgtg tggtcggtgc
     1861 tctggctggc tgtctctgct ccagggaagg cagcctcggg aaccttctgg tggctaggga
     1921 gcggctggta ccaatttgtt actttgattt cttggctgaa tgtctttctt cttaccaggt
     1981 gccttcgaaa tatttgcaag gtttttgtct tgctcctccc actcctactt ttactaggtg
     2041 ctggtgtctc cctgtggggc cagggaaact tcttctcact cctaccagtg ctgaactgga
     2101 cggccatgca gccaacacag agggtggacg attccaaggg catgcataga cctggccctc
     2161 ttcccccgag cccacctcca aaggttgatc acaaggcttc ccagtggcct caggagagtg
     2221 acatggggca gaaggtagct tctttgagtg cgcagtgcca caaccatgat gagagacttg
     2281 cagagctgac agtcctgctt cagaaactac agatacgggt agaccaagtg gatgacggca
     2341 gggaagggct gtcactgtgg gtcaagaatg tggttggaca gcacctgcag gagatgggca
     2401 ccatagaacc acctgatgct aagactgact tcatgacttt ccaccatgac catgaagtgc
     2461 gtctctccaa cttggaagat gttcttagaa aactgacaga aaaatctgag gctatccaga
     2521 aggagctgga agaaaccaag ctgaaagcag gcagcaggga tgaagagcag cccctccttg
     2581 accgtgtgca gcacctagaa ctggaactga acctgttgaa gtcacagctg tcagactggc
     2641 agcatctgaa gaccagctgt gagcaggctg gggcccgcat ccaggagact gtgcagctca
     2701 tgttctctga ggatcagcag ggcggttccc tcgagtggct attagagaag ctttcttctc
     2761 ggttcgtgag caaggatgag ctgcaggtgc tcttacatga ccttgagctg aaactgctgc
     2821 agaatatcac acaccacatc accgtgacag gacaggcccc gacatccgag gctattgtgt
     2881 ctgccgtgaa tcaggcaggg atttcaggaa tcacagaagc gcaagcacat atcattgtga
     2941 acaatgctct gaagctgtac tcccaagaca agacggggat ggtggacttt gctctggagt
     3001 ctggaggtgg cagcatccta agcactcggt gctctgagac ctatgagacc aagacggcac
     3061 tgctgagcct gtttggggtc ccactgtggt acttctcaca gtcacctcga gtggtgatcc
     3121 agcccgacat ctacccaggg aattgctggg cgttcaaagg ttcccagggg tacctggtgg
     3181 tgcggttgtc catgaagatc tacccaacca cattcaccat ggaacacatt ccaaagacac
     3241 tatcacccac tggtaacatc tccagtgccc ccaaagactt tgcagtctat ggactggaaa
     3301 cggagtatca agaagagggg cagcctctgg gacggttcac ctatgaccag gaaggagact
     3361 cactccagat gttccacaca ctggaaagac ctgaccaagc cttccagata gtagagctcc
     3421 gggtcctgtc caactggggc caccctgagt acacttgcct ctaccggttc cgagtccacg
     3481 gagagcccat ccagggaggc gtggcgcgcc tcgccgcagc ctccggcggc ggaggctctg
     3541 gcggaggggg ctccctcgca gcagcctctg aattcatggt gagcaagggc gaggagctgt
     3601 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
     3661 gcgtgcgcgg cgagggcgag ggcgatgcca ccaacggcaa gctgaccctg aagttcatct
     3721 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
     3781 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
     3841 tgcccgaagg ctacgtccag gagcgcacca tcagcttcaa ggacgacggc acctacaaga
     3901 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     3961 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tttaacagcc
     4021 acaacgtcta tatcaccgcc gacaagcaga agaacggcat caaggccaac ttcaagatcc
     4081 gccacaacgt ggaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     4141 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccgtgctga
     4201 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     4261 ggatcactct cggcatggac gagctgtaca agggaggcgg aggtgcaaga tctatggtga
     4321 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     4381 taaacggcca caagttcagc gtgcgcggcg agggcgaggg cgatgccacc aacggcaagc
     4441 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     4501 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
     4561 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc agcttcaagg
     4621 acgacggcac ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     4681 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     4741 agtacaactt taacagccac aacgtctata tcaccgccga caagcagaag aacggcatca
     4801 aggccaactt caagatccgc cacaacgtgg aggacggcag cgtgcagctc gccgaccact
     4861 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     4921 gcacccagtc cgtgctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     4981 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag agatctgaat
     5041 tcggcggagg atcccccggg ggccgctctg agcaaaagct catttctgaa gaggacttga
     5101 atggagaaca gaaattgatc agtgaggaag acctcaacgg tgagcagaag ttaatatccg
     5161 aggaggatct taatagtagc ggcagtggta gccgctctga gcaaaagctc atttctgaag
     5221 aggacttgaa tggagaacag aaattgatca gtgaggaaga cctcaacggt gagcagaagt
     5281 taatatccga ggaggatctt aatagtagcg gttaataagc ttcctcgact gtgccttcta
     5341 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     5401 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     5461 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     5521 gcaggcatgc tggggatgcg gtgggctcta tggcttctga gcataacgcg cttaattaag
     5581 gggatccggg atatcgggtc tatttaaatt ctgggggcct ttaaggccgc gcgcgtctag
     5641 agtctggtac cgggcctagg gggctcgagg ggtctcccag ctttcttgta caaagttggc
     5701 attataagaa agcattgctt atcaatttgt tgcaacgaac aggtcactat cagtcaaaat
     5761 aaaatcatta tttgccatcc agctgatatc ccctatagtg agtcgtatta catggtcata
     5821 gctgtttcct ggcagctctg gcccgtgtct caaaatctct gatgttacat tgcacaagat
     5881 aaaaatatat catcatgaac aataaaactg tctgcttaca taaacagtaa tacaaggggt
     5941 gttatgagcc atattcaacg ggaaacgtcg aggccgcgat taaattccaa catggatgct
     6001 gatttatatg ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat
     6061 cgcttgtatg ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt
     6121 gccaatgatg ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt
     6181 ccgaccatca agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc
     6241 cccggaaaaa cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt
     6301 gatgcgctgg cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt
     6361 aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt
     6421 gatgcgagtg attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa
     6481 atgcataaac ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt
     6541 gataacctta tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga
     6601 atcgcagacc gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct
     6661 tcattacaga aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg
     6721 cagtttcatt tgatgctcga tgagtttttc taatcagaat tggttaattg gttgtaacac
     6781 tggcagagca ttacgctgac ttgacgggac ggcgcaagct catgaccaaa atcccttaac
     6841 gtgagttacg cgtcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     6901 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     6961 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     7021 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     7081 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     7141 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     7201 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     7261 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     7321 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     7381 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     7441 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     7501 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
//
