LOCUS       sequence-220893-        7913 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology arms and linker-mEGFP sequence for C-terminus tagging of 
            human MYL2, via Cas9-excisable CAGGS-mCherry selection cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7913)
  AUTHORS   Roberts B, Hendershott MC, Arakaki J, Gerbin KA, Malik H, Nelson A, 
            Gehring J, Hookway C, Ludmann SA, Yang R, Haupt A, Grancharova T, 
            Valencia V, Fuqua MA, Tucker A, Rafelski SM, Gunawardane RN
  TITLE     Fluorescent Gene Tagging of Transcriptionally Silent Genes in 
            hiPSCs.
  JOURNAL   Stem Cell Reports. 2019 May 14;12(5):1145-1158. doi: 
            10.1016/j.stemcr.2019.03.001. Epub 2019 Apr 4.
  PUBMED    30956114
REFERENCE   2  (bases 1 to 7913)
  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: 
            "10.1016/j.stemcr.2019.03.001"; journalName: "Stem Cell Reports"; 
            date: "2019-05-14- 14"; volume: "12"; issue: "5"; pages: "1145-1158"
COMMENT     Sequence Label: AICSDP-49: MYL2-mEGFP
FEATURES             Location/Qualifiers
     source          1..7913
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1088..1209
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1125..1144)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1179..1198
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     enhancer        1224..1603
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1605..1882
                     /label=chicken beta-actin promoter
     intron          1884..2901
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2825..2847
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2909..2928
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(3129..3147)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3397..3416
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3702..3926
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             join(3995..3997,3998..4711)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4037..4058)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4298..4317)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4645..4666
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(join(5740..5746,5747..5764,5765..5770))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5785..5806
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5923..5940)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6094..6682)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6174..6193)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6853..7644,7645..7713))
                     /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     7476..7495
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7714..7818)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 yyrmrmgkky ygsykwyrcy ttccacctgc tgtgtgcttt gaacaagctc cttaacctca
       61 ctgcacctca atttcctcat ctgtaaagtg ggaataataa tagaagctac ctcataggag
      121 attgcgagga ttcgacatgt taatgtgggg aaagtgtgaa cttgtggcaa gggcaatata
      181 agtgctagct atgaatagcc agtatgtttt aaaaaaataa taataataac tagccaatgc
      241 tttttaaatg gggaaatttt gtgtaaaaac aaaaatctag atttctgaca ccctggaccc
      301 tgtggtcact tcatgtcacc tttttagatg gccacgtgtc tccatttagc cataggacct
      361 atcctgctcc cttgcataac ctgcctggcc cctcggaacc cttgagtttg tgatcccagt
      421 ggaaggctgg gtttccaggg acatctctgc atgatgttta cttgggaggg cccttggggt
      481 cagggaagga agaacaggga agggcaagaa aggaagaaca gggaagggca aggaaggaaa
      541 agtgcatgaa ggaagaagtt gagccaagat ttaggcccaa agatcggcac tgctgactac
      601 ccagggaggg agcttattct acagctctcc tgagaggtac caagttgggc gagatggccg
      661 tgcctttata cccccacacc aatcccatcc gctggctgtg ggctgcccca gaaagggcat
      721 gaccttgggc gagggggccc tgcagcagct ggggaagtcc ttccccggag gaccctctga
      781 gtggcccatg ttcccatccc ccacccccca gctatccgtc cactcaggcc ccgcccacag
      841 ccccccaccc tccgtctcag ttcccctccc ctcgtcctta gcacgtgttg ctggctcatt
      901 gcaggttgac cagatgttcg ccgccttccc ccctgacgtg actggcaacc tcgactacaa
      961 gaacctcgtg cacatcatca cacacggaga agagaaggac tccgggtccg ggtcccccgg
     1021 agaggttccc gacataccaa taacttcgta tagtgtagga tatacgaagt tatattagta
     1081 ttccgataac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     1141 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     1201 tgtatcttat catgtctggt ctcgacattg attattgact agttattaat agtaatcaat
     1261 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     1321 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     1381 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     1441 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     1501 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     1561 tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac
     1621 gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat
     1681 tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg cgccrggsgg
     1741 ggsggggsgg ggsgrggggs ggggsggggs gaggcggaga ggtgcggcgg cagccaatca
     1801 gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc ggccctataa
     1861 aaagcgaagc gcgcggcggg cggggagtcg ctgcgacgct gccttcgccc cgtgccccgc
     1921 tccgccgccg cctcgcgccg cccgccccgg ctctgactga ccgcgttact cccacaggtg
     1981 agcgggcggg acggcccttc tcctccgggc tgtaattagc gcttggttta atgacggctt
     2041 gtttcttttc tgtggctgcg tgaaagcctt gaggggctcc gggagggccc tttgtgcggg
     2101 gggagcggct cggggggtgc gtgcgtgtgt gtgtgcgtgg ggagcgccgc gtgcggctcc
     2161 gcgctgcccg gcggctgtga gcgctgcggg cgcggcgcgg ggctttgtgc gctccgcagt
     2221 gtgcgcgagg ggagcgcggc cgggggcggt gccccgcggt gcgggggggg ctgcgagggg
     2281 aacaaaggct gcgtgcgggg tgtgtgcgtg ggggggtgag cagggggtgt gggcgcgtcg
     2341 gtcgggctgc aaccccccct gcacccccct ccccgagttg ctgagcacgg cccggcttcg
     2401 ggtgcggggc tccgtacggg gcgtggcgcg gggctcgccg tgccgggcgg ggggtggcgg
     2461 caggtggggg tgccgggcgg ggcggggccg cctcgggccg gggagggctc gggggagggg
     2521 cgcggcggcc cccggagcgc cggcggctgt cgaggcgcgg cgagccgcag ccattgcctt
     2581 ttatggtaat cgtgcgagag ggcgcaggga cttcctttgt cccaaatctg tgcggagccg
     2641 aaatctggga ggcgccgccg caccccctct agcgggcgcg gggcgaagcg gtgcggcgcc
     2701 ggcaggaagg aaatgggcgg ggagggcctt cgtgcgtcgc cgcgccgccg tccccttctc
     2761 cctctccagc ctcggggctg tccgcggggg gacggctgcc ttcggggggg acggggcagg
     2821 gcggggttcg gcttctggcg tgtgaccggc ggctctagag cctctgctaa ccatgttcat
     2881 gccttcttct ttttcctaca gctcctgggc aacgtgctgg ttattgtgct gtctcatcat
     2941 tttggcaaag aattccgcca ccggggatcc accggagctt accatggtct ccaaaggaga
     3001 agaagataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc
     3061 cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac
     3121 ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct
     3181 gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg acatccccga
     3241 ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga
     3301 cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa
     3361 ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga agaagaccat
     3421 gggctgggag gcctcctccg agcggatgta ccccgaggac ggcgccctga agggcgagat
     3481 caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta
     3541 caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcaacatca agttggacat
     3601 cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca
     3661 ctccaccggc ggaatggatg aactctataa atgaggtact cctgtgcctt ctagttgcca
     3721 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     3781 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     3841 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     3901 tgctggggat gcggtgggct ctatggaaat aacttcgtat agtgtaggat atacgaagtt
     3961 atataggtat gtcgggaacc tctccgggtc cggggtgagc aagggcgagg agctgttcac
     4021 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     4081 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
     4141 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
     4201 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     4261 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     4321 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     4381 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     4441 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     4501 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     4561 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtcca agctgagcaa
     4621 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     4681 cactctcggc atggacgagc tgtacaagta ggagggggct cgctgctgcg cgctgggctc
     4741 gtctttgcag agtggtccct gccctcatct ctctcccccg agtaccgcct ctgtccctac
     4801 cttgtctgtt agccatgtgg ctgccccatt tatccacctc catcttcttt gcagcctggg
     4861 tggctatggg tacttcgtgg ccgcacatcc tacagttgga aatccatcca gaggccatgt
     4921 tccaataaac aggaggtcgt gtatttggtc acgacatttc tctgacaaac agtgccttgt
     4981 gtaaggaagg caaaggcaga tgagctagtg acagccttgc agtcattatt attgaatctt
     5041 ggtgactctt tatttaaggt agacctttgc agccactcag aactgtttct atcaaggatc
     5101 acgttttcct ggaatgttac ctggatgacc aaagccctct cttccaaaga gctcaagcat
     5161 ttagagggcc ttgcttccat gacatctgtc ccttgggatg ccatcggaat gcctctcctg
     5221 ccagtgacag tgacaagaag gaagcggact gacacatggt aggaacatag ctcgttcaac
     5281 agtaatcaca cctgctaatc ctaagagggt gacttctagt ttcagataaa gtagaagttg
     5341 tctgtacttc ttccctagca aactgaggga gtggtactga cttctgtctc tgtctggggc
     5401 ccaggcctgg atgtgagttg acctttgatt ttgaatgacc acagtcacgg ttgaagcatt
     5461 caggtctgtt atggctgcaa gagtcccagt gtaagaaaca aatagggcca ggtgtggtgg
     5521 ctcacgcctg tgatgctagc gctttgagag gctgaggcag gaggatcact tgagcccaaa
     5581 agtttgagac cagcctaggc aacatagtga gatccaatct ctataaaaaa taaatagcca
     5641 ggcatggtgg tgtgcacctg tggtctcagc tacccaggag gctgcagtgg gaagatcgct
     5701 tgwgccyrwk ysmaatkgyk mkccgmkcac aatkccacac aacatacgag ccggaagcat
     5761 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     5821 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5881 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5941 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     6001 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     6061 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     6121 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     6181 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     6241 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     6301 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6361 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6421 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6481 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     6541 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6601 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6661 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6721 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6781 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6841 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6901 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6961 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     7021 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     7081 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     7141 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     7201 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     7261 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     7321 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7381 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7441 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7501 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7561 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7621 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7681 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7741 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7801 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     7861 tattatcatg acattaacct ataaaaatag gcgtatcacg aggsymttks gwy
//
