LOCUS       sequence-177429-        5418 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology arms and linker-mEGFP sequence for N-terminus tagging of 
            human MYH10.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5418)
  AUTHORS   Roberts B, Haupt A, Tucker A, Grancharova T, Arakaki J, Fuqua MA, 
            Nelson A, Hookway C, Ludmann SA, Mueller IA, Yang R, Horwitz R, 
            Rafelski SM, Gunawardane RN
  TITLE     Systematic gene tagging using CRISPR/Cas9 in human stem cells to 
            illuminate cell organization.
  JOURNAL   Mol Biol Cell. 2017 Oct 15;28(21):2854-2874. doi: 
            10.1091/mbc.E17-03-0209. Epub 2017 Aug 16.
  PUBMED    28814507
REFERENCE   2  (bases 1 to 5418)
  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: 
            "10.1091/mbc.E17-03-0209"; journalName: "Mol Biol Cell"; date: 
            "2017-10-15- 15"; volume: "28"; issue: "21"; pages: "2854-2874"
COMMENT     Sequence Label: AICSDP-24:MYH10-mEGFP
FEATURES             Location/Qualifiers
     source          1..5418
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             join(1413..1415,1416..1418,1419..2129)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1458..1479)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1719..1738)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2066..2087
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3197..3213)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3197..3213)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3210..3232)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3221..3237
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3245..3251,3252..3269,3270..3275))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3290..3311
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3428..3445)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3599..4187)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3679..3698)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4358..5149,5150..5218))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4981..5000
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5219..5323)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5391..5409
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gacgcgtatt gggataagtg
      421 atggagcttc ctctgctagc cctttgtgag ccaatggtaa atgggtgcta aataaaacaa
      481 ctaggtcttg agatacatta attgtaaatg tcacagaacc agtactttcc tcaatgtggc
      541 taagatagtt gatggttcct ttttcttctg cactggtcag accatatctg ggctatgatg
      601 tttgcttctg ggccacacac tttagaggga agacaagcag catgttggag tctgtttagg
      661 cgagagatcc ggcaggggaa ggagtcttgg tgaatgaggg gtggaggagc tgcaggatgg
      721 gaatagaggc ctgaactgct accatgacat attcaaaagg ctgccgtgtg aagccaagtt
      781 atgcttgtct tttgtggtcc cagttgatca cattaagacc tcatggggcc atagaaaagc
      841 tagagggaga ctgatttggg ttattcataa gaagaacttt aagtctgtta tctgagggta
      901 gaatgagagg catgttttag attctttaga ttctttactc ttctgacaat catgtgtttt
      961 tgtagctgtt tccttgtggt catattaatt ctggtaccac ttcatgaacc ttttattacc
     1021 catctttgtt ttcttttttt ttttcccttc ttaactccct gtttaatttg tggtgagggt
     1081 gaaagaggag ataaagaaaa aaaagggtca acttgtaact ttgccttttc ttttcttttc
     1141 ttttcttttt tttgccctca gtaactgagg gcaaacccat cagacaacca gagccataat
     1201 ttgtggtcac cgctgaaatt taccttggaa actcggttag tatggctgtg aagaggtata
     1261 ccccagctcc ttaacacaga gttaatgctt aatctaaggt tttaagtttc ttagaaaaga
     1321 aaaacgtgta cattcttttg tttcttaaac atctaatttt gccctcctcc tcttctctta
     1381 gaggcaattg cttttggatc gttccattta caatggtgag caagggcgag gagctgttca
     1441 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     1501 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     1561 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     1621 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     1681 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     1741 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     1801 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     1861 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     1921 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     1981 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc aagctgagca
     2041 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     2101 tcactctcgg catggacgag ctgtacaagt acagcgatct ggagctgaag ctgcggatcc
     2161 ctgcacaaag gacagggctg gaggatccag agaggtacct gttcgtcgat cgtgctgtca
     2221 tctacaaccc tgccactcaa gctgattgga cagctaaaaa gctagtgtgg attccatcag
     2281 aacgccatgg ttttgaggca gctagtatca aagaagaacg gggagatgaa gttatggtgg
     2341 agttggcaga gaatggaaag aaagcaatgg tcaacaaaga tgatattcag aagatgaacc
     2401 cacctaagtt ttccaaggtg gaggatatgg cagaattgac atgcttgaat gaagcttccg
     2461 ttttacataa tctgaaggat cgctactatt caggactaat ctatgtaagt atttcttcca
     2521 aataatcatg tgaagtggta gctaggaatt aatgtaaatt atacatcttg tcataatcaa
     2581 atgagaatgt ggaataccca aactctctgt ttaacatttc tatttctctt taagatagaa
     2641 agatttgttg cttgcttacc catgtcttgc ttttctttga atcttaacac attaagttta
     2701 aataatacag gctgcaatta catataataa aatggcattt gaagactttt gtagtggtct
     2761 tctggagcat aataaggtgg gagagagcat gtaacaggaa gaccagaagg tttaataagg
     2821 taaagagagt tgcattaatt ggacgcagac agcaaaacgg tcaaaaatca agtgcatacc
     2881 caagagtaaa gtggaggggc tgtaagctga gaaatttctg tggacagcat gaacagcttc
     2941 actggatgta gtagggaagt aggaaagatg aatgctgagg tttttaagag gaaacaatta
     3001 gggtaagatt gaggctggct ggggtcgtcc tgtggttagc agctgacatg aatgttggag
     3061 tcaccgactt tgtcactgac catgtagaag aagttattga aaatcataag ggataatgta
     3121 gagagggata atgtagagag gaaaaatgta agccagatac taatcccaat ggcgcgccga
     3181 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     3241 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     3301 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     3361 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     3421 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     3481 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     3541 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     3601 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3661 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3721 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     3781 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     3841 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     3901 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     3961 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4021 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4081 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4141 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4201 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4261 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     4321 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     4381 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4441 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     4501 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     4561 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     4621 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     4681 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     4741 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     4801 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     4861 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     4921 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     4981 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5041 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     5101 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5161 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     5221 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     5281 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     5341 tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta
     5401 tcacgaggcc ctttcgtc
//
