LOCUS       sequence-377386-        7356 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses EGFP-tagged MASTL with E167D mutation and resistance to 
            MASTL siRNA (ACGCCTTATTCTAGCAAATTA).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7356)
  AUTHORS   Taskinen ME, Narva E, Conway JRW, Hinojosa LS, Lilla S, Mai A, De 
            Franceschi N, Elo LL, Grosse R, Zanivan S, Norman JC, Ivaska J
  TITLE     MASTL promotes cell contractility and motility through 
            kinase-independent signaling.
  JOURNAL   J Cell Biol. 2020 Jun 1;219(6). pii: 151688. doi: 
            10.1083/jcb.201906204.
  PUBMED    32311005
REFERENCE   2  (bases 1 to 7356)
  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: "J Cell 
            Biol."; date: "2020-06-1"; volume: "219(6). pii: 151688. doi"; 
            pages: " 10.1083/jcb.201906204"
COMMENT     Sequence Label: pcDNA-EGFP MASTL E167D (siRNA resistant)
FEATURES             Location/Qualifiers
     source          1..7356
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        69..372
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        373..576
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     527..547
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      615..624
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(621..623,624..626,627..1337)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(621..623,624..626,627..1337)
                     /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(666..687)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(927..946)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1274..1295
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4152..4273
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4189..4208)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4243..4262
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4280..4735)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4417..4438)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4629..4648
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4762..4866
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4868..5225
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4868..5059
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4891..4911)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5076..5211
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5260..6054
                     /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             5260..6054
                     /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(5314..5333)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5924..5943
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6242..6261)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6286..6333
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6662..7250
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7151..7170
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccatgcatta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
       61 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
      121 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      181 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      241 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      301 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      361 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      421 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      481 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      541 aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg tcagatccgc
      601 tagcgctacc ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
      661 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
      721 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
      781 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
      841 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
      901 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
      961 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     1021 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     1081 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     1141 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     1201 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     1261 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     1321 tggacgagct gtacaagtcc ggactcagat ctcgagctca agcttccatg gatcccaccg
     1381 cgggaagcaa gaaggagcct ggaggaggcg cggcgactga ggagggcgtg aataggatcg
     1441 cagtgccaaa accgccctcc attgaggaat tcagcatagt gaagcccatt agccggggcg
     1501 ccttcgggaa agtgtatctg gggcagaaag gcggcaaatt gtatgcagta aaggttgtta
     1561 aaaaagcaga catgatcaac aaaaatatga ctcatcaggt ccaagctgag agagatgcac
     1621 tggcactaag caaaagccca ttcattgtcc atttgtatta ttcactgcag tctgcaaaca
     1681 atgtctactt ggtaatggaa tatcttattg ggggagatgt caagtctctc ctacatatat
     1741 atggttattt tgatgaagag atggctgtga aatatatttc tgaagtagca ctggctctag
     1801 actaccttca cagacatgga atcatccaca gggacttgaa accggacaat atgcttattt
     1861 ctaatgacgg tcatattaaa ctgacggatt ttggcctttc aaaagttact ttgaatagag
     1921 atattaatat gatggatatc cttacaacac catcaatggc aaaacctaga caagattatt
     1981 caagaacccc aggacaagtg ttatcgctta tcagctcgtt gggatttaac acaccaattg
     2041 cagaaaaaaa tcaagaccct gcaaacatcc tttcagcctg tctgtctgaa acatcacagc
     2101 tttctcaagg actcgtatgc cctatgtctg tagatcaaaa ggacactacc ccctacagct
     2161 ctaagctgct aaaatcatgt cttgaaacag ttgcctccaa cccaggaatg cctgtgaagt
     2221 gtctaacttc taatttactc cagtctagga aaaggctggc cacatccagt gccagtagtc
     2281 aatcccacac cttcatatcc agtgtggaat cagaatgcca cagcagtccc aaatgggaaa
     2341 aagattgcca ggaaagtgat gaagcattgg gcccaacaat gatgagttgg aatgcagttg
     2401 aaaagttatg cgcaaaatct gcaaatgcca ttgagacgaa aggtttcaat aaaaaggatc
     2461 tggagttagc tctttctccc attcataaca gcagtgccct tcccaccact ggacgctctt
     2521 gtgtaaacct tgctaaaaaa tgcttctctg gggaagtttc ttgggaagca gtagaactgg
     2581 atgtaaataa tataaatatg gacactgaca caagtcagtt aggtttccat cagtcaaatc
     2641 agtggactgt ggattctggt gggatatctg aagagcacct tgggaaaaga agtttaaaaa
     2701 gaaattttga gttggttgac tccagtcctt gtaaaaaaat tatacagaat aaaaaaactt
     2761 gtgtagagta taagcataac gaaatgacaa attgttatac aaatcaaaat acaggcttaa
     2821 cagttgaagt gcaggacctt aagctatcag tgcacaaaag tcaacaaaat gactgtgcta
     2881 ataaggagaa cattgtcaat tcttttactg ataaacaaca aacaccagaa aaattaccta
     2941 taccaatgat agcaaaaaac cttatgtgtg aactcgatga agactgtgaa aagaatagta
     3001 agagggacta cttaagttct agttttctat gttctgatga tgatagagct tctaaaaata
     3061 tttctatgaa ctctgattca tcttttcctg gaatttctat aatggaaagt ccattagaaa
     3121 gtcagccctt agattcagat agaagcatca aagaatcctc ttttgaagaa tcaaatattg
     3181 aagatccact tattgtaaca ccagattgcc aagaaaagac ctcaccaaaa ggtgtcgaga
     3241 accctgctgt acaagagagt aaccaaaaaa tgttaggtcc tcctttggag gtgctgaaaa
     3301 cgttagcctc taaaagaaat gctgttgctt ttcgaagttt taacagtcat attaatgcat
     3361 ccaataactc agaaccatcc agaatgaaca tgacttcttt agatgcaatg gatatttcgt
     3421 gtgcctacag tggttcatat cccatggcta taacccctac tcaaaaaaga agatcctgta
     3481 tgccacatca gcagacccca aatcagatca agtcgggaac tccataccga actccgaaga
     3541 gtgtgagaag aggggtggcc cccgttgatg atgggcgaat tctaggaacc ccagactacc
     3601 ttgcacctga gctgttacta ggcagggccc atggtcctgc ggtagactgg tgggcacttg
     3661 gagtttgctt gtttgaattt ctaacaggaa ttcccccttt caatgatgaa acaccacaac
     3721 aagtattcca gaatattctg aaaagagata tcccttggcc agaaggtgaa gaaaagttat
     3781 ctgataatgc tcaaagtgca gtagaaatac ttttaaccat tgatgataca aagagagctg
     3841 gaatgaaaga gctaaaacgt catcctctct tcagtgatgt ggactgggaa aatctgcagc
     3901 atcagactat gcctttcatc ccccagccag atgatgaaac agatacctcc tattttgaag
     3961 ccaggaatac tgctcagcac ctgaccgtat ctggatttag tctgtagggt accgcgggcc
     4021 cgggatccac cggatctaga taactgatca taatcagcca taccacattt gtagaggttt
     4081 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     4141 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     4201 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4261 tcaatgtatc ttaacgcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     4321 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     4381 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     4441 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     4501 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     4561 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     4621 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     4681 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt
     4741 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     4801 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     4861 aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     4921 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4981 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     5041 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     5101 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     5161 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     5221 gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     5281 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     5341 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     5401 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta
     5461 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     5521 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     5581 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     5641 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     5701 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     5761 gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc
     5821 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     5881 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     5941 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     6001 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     6061 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     6121 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     6181 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa
     6241 ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac
     6301 agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct
     6361 ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct
     6421 tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg
     6481 cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc
     6541 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     6601 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     6661 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     6721 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     6781 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     6841 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     6901 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     6961 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     7021 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     7081 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     7141 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     7201 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     7261 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     7321 cgttatcccc tgattctgtg gataaccgta ttaccg
//
