LOCUS       sequence-248752-        6907 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses EGFP-deltaN-term-MCAK in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6907)
  AUTHORS   Domnitz SB, Wagenbach M, Decarreau J, Wordeman L
  TITLE     MCAK activity at microtubule tips regulates spindle microtubule 
            length to promote robust kinetochore attachment.
  JOURNAL   J Cell Biol. 2012 Apr 16;197(2):231-7. doi: 10.1083/jcb.201108147. 
            Epub 2012 Apr 9.
  PUBMED    22492725
REFERENCE   2  (bases 1 to 6907)
  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: 
            "10.1083/jcb.201108147"; journalName: "J Cell Biol"; date: 
            "2012-04-16- 16"; volume: "197"; issue: "2"; pages: "231-7"
COMMENT     Sequence Label: pMX1433
FEATURES             Location/Qualifiers
     source          1..6907
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3493..3510
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    3695..3816
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3732..3751)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3786..3805
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3823..4278)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3960..3981)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4172..4191
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4305..4409
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4411..4768
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4411..4602
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4434..4454)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4619..4754
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4803..5597
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4803..5597
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4857..4876)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5467..5486
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5785..5804)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5829..5876
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6205..6793
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6694..6713
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggaccggt gctagcggta ccgagctcgg atccactagt ccagtgtggt
     1381 ggaattgccc ttgccaatgg cttaattcac agtgccaaca taagcacagt gaacgtggag
     1441 aaatcctgtg tttcagtgga atggatagaa ggaggtaaca cgaaaggcaa agagattgat
     1501 tttgatgatg tggctgccat caatccggag cttctgcagc ttcttccctt acacccaaag
     1561 gacaatctgc cactgcagga aaatgtaaca gttccgaagc aaaaacgcag atcagtcaac
     1621 tccaaaattc cagctccaaa agaaggtctt cgaagccgtt ccactcgcat gtccactgtc
     1681 ccagaggttc gaattgccac tcaggagaat gaaatggagg tggagctgcc tgtggctaca
     1741 aactctcgaa agcagttttc ggttgccact ggcctcccta ggccctcctg tcctgcaatg
     1801 acagaattac cgttgtcgat ggtcagcgag gaggcagaag agcaagtcca tcccacccga
     1861 agcacatctt ctgcaaaccc tgctcggagg aaatcatgta ttgtgaagga aatggaaaaa
     1921 atgaagaaca agcgggaaga gaagagggcc cagaactcgg aaataagaat aaagcgagct
     1981 caggaatatg acagcagctt tccaaactgg gaatttgccc ggatgattaa agaattccgg
     2041 gttactatag aatgtcaccc acttactctg actgatccta cggaagagca caggatctgc
     2101 gtgtgtgtga ggaagcgccc actgaataag caagaactgg ccaagaaaga aatcgatgtg
     2161 atttctgttc ccagcaagtg tttgctcttt gtgcatgaac ccaaattgaa agtggacttg
     2221 acaaagtatc tggagaatca ggcattctgc ttcgattttg cctttgatga aacagcttca
     2281 aatgaagtcg tctacaggtt cacagcaagg ccactggtac agacgatttt tgaaggggga
     2341 aaagcaacct gctttgccta tgggcagacg ggcagtggga agactcacac aatgggtgga
     2401 gacctgtctg gtaaatctca gaatacatct aaagggatct atgcaatggc ttcccgggat
     2461 gtcttccttc tgaagagtca gcctcgctac cggaacttaa acttggaagt ttatgtgaca
     2521 ttctttgaga tctacaatgg gaaggtgttt gatctgctca acaagaaggc caagctacgt
     2581 gtactagaag acagcaagca gcaggtgcag gttgtgggac ttcaggagta cctggttaac
     2641 tgtgctgatg atgtcatcaa gatgctcaac atgggcagtg cctgcaggac ttctggacag
     2701 acttttgcca actccaattc ttcccgctcc catgcctgct tccagattct tcttcgagcc
     2761 aaagggagat tgcatggcaa attctccttg gtggatctgg cagggaatga acgaggggct
     2821 gacacttcta gtgctgaccg ccagactcgc atggagggtg cagagattaa caagtctctc
     2881 ctagcactga aggagtgcat cagggcgctg ggacagaaca aggctcacac cccattccgt
     2941 gagagcaagc tgactcaggt gctaagggat tcattcattg gggagaactc gaggacttgc
     3001 atgattgcca tgatctcacc aggcataagc tcctgtgaat atactttaaa cacgctgaga
     3061 tatgcagaca gagtcaagga gctgagcccc cacagcgggc tcagtggaga gcagccaatt
     3121 caaatggaaa cggaagagat ggaagccagc tctaatggga cctcgctggc agtcaatttc
     3181 aaggaagaag aggagctgtc ttcccagatg tccagcttta atgaagccat gagtcagatc
     3241 agggagctgg aagagagggc catggaggag ctcagggaga tcatacagca aggaccaggc
     3301 tggcttgagc tctctgagat gactgatcag ccagactatg acttagagac ctttgtgaac
     3361 aaagcagaat ctgccctgac tcagcagact aagcacttct cagccctgcg agaagtcatc
     3421 aaggccctgc gtgtggccat gcaactggaa gaacaggcta gcaaacagat gaacagcaag
     3481 aaacggcacc aacaccatca ccatcatcac taagcggccg catcgataag cttcgaattc
     3541 tgcagtcgac ggtaccgcgg gcccgggatc caccggatct agataactga tcataatcag
     3601 ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa
     3661 cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg
     3721 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     3781 tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta agcgttaata
     3841 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     3901 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     3961 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     4021 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     4081 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     4141 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     4201 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     4261 cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     4321 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     4381 tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt
     4441 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     4501 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     4561 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     4621 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     4681 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     4741 cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc
     4801 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     4861 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     4921 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     4981 tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     5041 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     5101 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     5161 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     5221 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     5281 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg
     5341 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     5401 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     5461 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     5521 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     5581 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     5641 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     5701 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     5761 cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg
     5821 cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa
     5881 acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg
     5941 ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc
     6001 agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat
     6061 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     6121 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     6181 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6241 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6301 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     6361 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     6421 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     6481 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     6541 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     6601 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     6661 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     6721 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     6781 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     6841 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     6901 catgcat
//
