LOCUS       sequence-250393-        5238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of plant nuclear envelope LINC protein fused to 
            fluorescent protein and affinity tags for cell biology.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5238)
  AUTHORS   Gumber HK, McKenna JF, Tolmie AF, Jalovec AM, Kartick AC, Graumann 
            K, Bass HW
  TITLE     MLKS2 is an ARM domain and F-actin-associated KASH protein that 
            functions in stomatal complex development and meiotic chromosome 
            segregation.
  JOURNAL   Nucleus. 2019 Dec;10(1):144-166. doi: 10.1080/19491034.2019.1629795.
  PUBMED    31221013
REFERENCE   2  (bases 1 to 5238)
  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"; doi: 
            "10.1080/19491034.2019"; journalName: "Nucleus"; date: "2019-12"; 
            volume: "10"; issue: "1"; pages: "144-166"
COMMENT     Sequence Label: MLKS2del4ec
FEATURES             Location/Qualifiers
     source          1..5238
                     /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             join(671..673,674..676,677..1387)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             1397..1420
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1424..1450
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    complement(3363..3462)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3479..3498)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3480..3498)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3503..3519)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3503..3519)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3621..3640)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3632..4441
                     /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(3699..3718)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4588..5176
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5077..5096
                     /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 agcaggcttc atggtctcca agggcgagga gctgttcacc ggggtggtcc ccatcctggt
      721 ggagctggat ggcgacgtta acgggcacaa gttctcggtg tctggcgagg gggagggcga
      781 cgctacctac gggaagctca cgctgaagtt catctgcacg accggcaagc tgccggtgcc
      841 ctggccaaca ctcgtcacaa ctctgactta cggcgtccag tgcttctcgc gctaccctga
      901 tcacatgaag cagcatgact tcttcaagtc ggctatgccc gagggctacg tgcaggagcg
      961 gacgatcttc ttcaaggacg atggcaacta caagacacgc gcggaggtca agttcgaggg
     1021 ggacactctc gttaacagga tcgagctgaa gggcattgac ttcaaggagg atggcaatat
     1081 cctcgggcac aagctggagt acaactacaa ttcccataac gtgtacatca tggccgacaa
     1141 gcagaagaac ggcattaagg tcaatttcaa gatccggcac aacattgagg atggcagcgt
     1201 tcagctcgcg gaccattacc agcagaacac accgatcggg gatggccctg tcctcctgcc
     1261 cgacaatcac tacctctcca ctcagagcgc cctgtcgaag gacccgaacg agaagaggga
     1321 ccatatggtg ctcctggagt tcgtcacagc cgcgggcatc actctcggga tggatgagct
     1381 gtacaagggg gcgggcgact acaaggacga tgacgataag gcttacccct acgacgtgcc
     1441 ggattacgcc ggggctggca tggggcgctc actctccccg ctcctgaggc aggagctgga
     1501 caacctggat aaggacgccg attctcgcag ggctgccatg aaggctctca agtcatacgc
     1561 ccgccacctg gatagcaagt cgatccccca tttcctggcc gaggtctctg acacgaccgc
     1621 cgctggcgcg ggggccggca gcgggatgcc aaccggcggg gagttcacga tctcgctcta
     1681 cgaggtcctg gccagggttc acgggcggaa catcgtgccc cagatcggca atattatggc
     1741 gaccattatg tgcacgctct ccagctcggg cgggtcattc ccactgcacc aggcctgctc
     1801 caaggttgtg ccagctatcg ctaggcatgg cattgatccg tcggccccag acggcgagaa
     1861 ggccgggatc attgctagcc tctgccgccc actgtgctcg gctctcatgg gcgggagcca
     1921 ggacggcggc gctgccagcg gcgctgccct ctgcctgaag gccctggttg agtcttcaaa
     1981 ctggaggttc gcgtcgggcg agctggtgaa tgaggtctgc ctgaaggtgg ccggcgctat
     2041 gcacgactcg gccacacggt cgaacgctca tatggggctg gccatggcgc tcgtgaagca
     2101 caatggcctc atcgctgagg cctacgcgcg ctctatcgtc aggtccggcc tccagattct
     2161 ggacggggat accgctgact ccagctcgca gaagaggctg tccgccatcc agatgattaa
     2221 cttcttcatg aagttcgtgg acccgaggtg cctgtcttca gagctgggcc gggttatcga
     2281 tgtgatggag cgctgccaga atgaccggat gcccttcgtg cggggcgcgg ctttcgaggc
     2341 ctctcagtca gcgcgcaaca ttgccgcgca gaagggctcc aggcacgagg ttgggacctc
     2401 tcctatggtg ggctcaaact tccataagag gagggagaag tccccttgcc ggagcctctg
     2461 gggcgccaag aggtcccccg ctgcctctac gatggccgct gctagccctg tgcagttccg
     2521 ctctccggag tcacaggtcg ttgattccag catcatgaat ggctccacac tccacgactc
     2581 cccagtgagc gtcggccagt cgtcttgcaa cttcgaccag agcaggcgga ctaaccgcag
     2641 gctgtggtcg aatgacgcgg ttgatgtgtc tctgaaggac ggcctcttca tccagctgtg
     2701 ctccaacagc aattcgtacg aggacgatct gggcgaggtt tgcgatagcg aggtgaccga
     2761 cgctaacttc gagtgcacca atacgttcgc cgggttcgtc tctccctcac acaatggctc
     2821 catcagcagg gataagacgc cttcaccgcg ggcctcggac aggcccatca acattgacga
     2881 tgtgaagatc tactccacgc caaggaagct cctgcggctc ctgcagtcat cctacgactc
     2941 ggattctgcc tcaaacatcg ggcagagcac ggccaagctc aggggcctct cctcccccga
     3001 ccaggagcac aagggcctcg tcatttcttc agagcagatg cagtcgctgc attctgacaa
     3061 taagggcgat gagatgaagg acgagaacga gaccatctac atgcagaata gcaagggcag
     3121 gacacagact ctctcgaacg ctgacgcctc agtcctgtcc acaactgagg ccgagaacac
     3181 aagcatcaag acttcgcctg agattgagct gaatggggac gatgtttgca tcacctccag
     3241 cattggcaag acgcggaagt accgcgcgaa gttcaccctc ttcctgtccg tcatcgttat
     3301 tttcctcgct atcattacga tgctgatccg cattgagaac gacgatgaca gcgcgtactg
     3361 agacccagct ttcttgtaca aagttggcat tataagaaag cattgcttat caatttgttg
     3421 caacgaacag gtcactatca gtcaaaataa aatcattatt tgccatccag ctgatatccc
     3481 ctatagtgag tcgtattaca tggtcatagc tgtttcctgg cagctctggc ccgtgtctca
     3541 aaatctctga tgttacattg cacaagataa aaatatatca tcatgaacaa taaaactgtc
     3601 tgcttacata aacagtaata caaggggtgt tatgagccat attcaacggg aaacgtcgag
     3661 gccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa
     3721 tgtcgggcaa tcaggtgcga caatctatcg cttgtatggg aagcccgatg cgccagagtt
     3781 gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact
     3841 aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga
     3901 tgatgcatgg ttactcacca ctgcgatccc cggaaaaaca gcattccagg tattagaaga
     3961 atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca
     4021 ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc
     4081 gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg
     4141 ctggcctgtt gaacaagtct ggaaagaaat gcataaactt ttgccattct caccggattc
     4201 agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat
     4261 aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct
     4321 atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg
     4381 tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta
     4441 atcagaattg gttaattggt tgtaacactg gcagagcatt acgctgactt gacgggacgg
     4501 cgcaagctca tgaccaaaat cccttaacgt gagttacgcg tcgttccact gagcgtcaga
     4561 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     4621 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     4681 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     4741 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     4801 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     4861 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     4921 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     4981 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     5041 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     5101 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     5161 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     5221 gccttttgct cacatgtt
//
