LOCUS       sequence-250980-        8376 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology arms and linker-mEGFP sequence for C-terminus tagging of 
            human MYL7, via Cas9-excisable CAGGS-mCherry selection cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8376)
  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 8376)
  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-46: MYL7-mEGFP
FEATURES             Location/Qualifiers
     source          1..8376
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    392..513
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(429..448)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     483..502
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     enhancer        528..907
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        909..1186
                     /label=chicken beta-actin promoter
     intron          1188..2205
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2129..2151
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2213..2232
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(2433..2451)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2701..2720
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3006..3230
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             join(3299..3301,3302..4015)
                     /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(3341..3362)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3602..3621)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3949..3970
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(5047..5063)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5047..5063)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5060..5082)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5071..5087
                     /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(5095..5101,5102..5119,5120..5125))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5140..5161
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5278..5295)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5449..6037)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5529..5548)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6208..6999,7000..7068))
                     /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     6831..6850
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7069..7173)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7241..7259
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7297..7319)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7419..7438
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7632..7654
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7646..7663
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7647..7663
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ccttggccca catttgcact gactcctcac tctgcccaga gtcagccaag agaaaaacat
       61 taacccagag tctggggtct agggttgaaa agctaaggca aaaagcacag atgcaggggg
      121 cagacagaaa ggccacagga ctcaggtgag gtctctgccg ggctgggcca ggagccaggg
      181 gactgccact caccagtgtc ccctgcaggt ggagcagatg ttcgccctga cacccatgga
      241 tttggcgggg aacatcgact acaagtcact gtgctacatt attacacatg gagacgagaa
      301 agaggagggg tccgggtccc ccggagaggt tcccgacata ccaataactt cgtatagtgt
      361 aggatatacg aagttatatt agtattccga taacttgttt attgcagctt ataatggtta
      421 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
      481 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggtctcgac attgattatt
      541 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
      601 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
      661 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
      721 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
      781 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
      841 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
      901 taccatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc
      961 acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg
     1021 gggggggggg cgcgcgccrg gsggggsggg gsggggsgrg gggsggggsg gggcgaggcg
     1081 gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
     1141 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggga gtcgctgcga
     1201 cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc ccggctctga
     1261 ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc gggctgtaat
     1321 tagcgcttgg tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag ccttgagggg
     1381 ctccgggagg gccctttgtg cggggggagc ggctcggggg gtgcgtgcgt gtgtgtgtgc
     1441 gtggggagcg ccgcgtgcgg ctccgcgctg cccggcggct gtgagcgctg cgggcgcggc
     1501 gcggggcttt gtgcgctccg cagtgtgcgc gaggggagcg cggccggggg cggtgccccg
     1561 cggtgcgggg ggggctgcga ggggaacaaa ggctgcgtgc ggggtgtgtg cgtggggggg
     1621 tgagcagggg gtgtgggcgc gtcggtcggg ctgcaacccc ccctgcaccc ccctccccga
     1681 gttgctgagc acggcccggc ttcgggtgcg gggctccgta cggggcgtgg cgcggggctc
     1741 gccgtgccgg gcggggggtg gcggcaggtg ggggtgccgg gcggggcggg gccgcctcgg
     1801 gccggggagg gctcggggga ggggcgcggc ggcccccgga gcgccggcgg ctgtcgaggc
     1861 gcggcgagcc gcagccattg ccttttatgg taatcgtgcg agagggcgca gggacttcct
     1921 ttgtcccaaa tctgtgcgga gccgaaatct gggaggcgcc gccgcacccc ctctagcggg
     1981 cgcggggcga agcggtgcgg cgccggcagg aaggaaatgg gcggggaggg ccttcgtgcg
     2041 tcgccgcgcc gccgtcccct tctccctctc cagcctcggg gctgtccgcg gggggacggc
     2101 tgccttcggg ggggacgggg cagggcgggg ttcggcttct ggcgtgtgac cggcggctct
     2161 agagcctctg ctaaccatgt tcatgccttc ttctttttcc tacagctcct gggcaacgtg
     2221 ctggttattg tgctgtctca tcattttggc aaagaattcc gccaccgggg atccaccgga
     2281 gcttaccatg gtctccaaag gagaagaaga taacatggcc atcatcaagg agttcatgcg
     2341 cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg
     2401 cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc
     2461 cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt
     2521 gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg
     2581 ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct
     2641 gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg
     2701 ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga
     2761 ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta
     2821 cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta
     2881 caacgtcaac atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca
     2941 gtacgaacgc gccgagggcc gccactccac cggcggaatg gatgaactct ataaatgagg
     3001 tactcctgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     3061 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     3121 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     3181 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg aaataacttc
     3241 gtatagtgta ggatatacga agttatatag gtatgtcggg aacctctccg ggtccggggt
     3301 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     3361 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     3421 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     3481 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     3541 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     3601 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     3661 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     3721 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     3781 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     3841 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     3901 gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     3961 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtgaggggc
     4021 agggccaggc ccacgggggg gcacctcaat aaactctgtt gcaaaattgg aattgctgtg
     4081 gtgtcttgtc tgtgacagat gcgttgggga ccagccaagg gggatcccag ggtctcagtg
     4141 cgcacatcac catgatcatg gccaccatct acctcctggg agctggcccc tcgccagctc
     4201 accttgattc actcccatga tgccaagtga agtgtgaact atgatcatgc ctagtttaca
     4261 gatgaggaca ctgaggccca gaaagtgtga gcatcttacc aaggccagcc ctctagaaga
     4321 ggagatggtg ggatttacac cacctccacc aagcccagga atgagccaca aagtgggcac
     4381 tgcccagcta cttggggctg tgcagagaag aggctgcttg ctgggcactc agcaaactct
     4441 gcccaacagc ccagcgggtg ggcagcagcc ctgggacccc cacacccaac cacacagcct
     4501 cccctggccc actgctcgca ccccatctca atacactggc ttgggtgcct ccctgcatgg
     4561 gccctttgtg aaaggcagag aggtacccat ttgaaacaca accagcttct cattgcaaat
     4621 acaggcaagg cactaagaca tgaggaacat ggacaccaaa gcaggggcca ggtaacatgc
     4681 aaatttctag aggaaatgcc cagaacctgg catcatgcct cctgagcccc tcatgcgccg
     4741 tgaggggtaa gagggtcaga cagctggagt gtagggagac gacttctcag gagagaatag
     4801 ttagtgctcc cgtcaccctt catctgagaa cccaagagct agaggagaaa gtgatcctca
     4861 tgagtaccag aggagcagca ggggacatcc aaagcaccag agagagaaac agagacagag
     4921 agacaggcag tgacagctca aacctcagcc agatccagag catacaaagt ctcctgccta
     4981 caggacagcc cagtaagagc tctcagcttg ccatcccaat ggcgcgccga gcttggcgta
     5041 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     5101 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     5161 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     5221 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     5281 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5341 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5401 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5461 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5521 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5581 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5641 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5701 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5761 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     5821 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5881 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5941 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     6001 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     6061 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6121 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     6181 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     6241 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     6301 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     6361 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     6421 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     6481 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     6541 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     6601 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     6661 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     6721 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     6781 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     6841 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     6901 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     6961 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     7021 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     7081 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     7141 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     7201 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     7261 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     7321 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     7381 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
     7441 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     7501 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     7561 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     7621 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgaattga cgcgtattgg
     7681 gatctcctgc gggggacctg ggatgctgga gacatggatg ctcacctggc tgcctcggcc
     7741 ttccagggac agaccccgag gaagccatcc tgagtgcctt ccgcatgttt gaccccagcg
     7801 gcaaaggggt ggtgaacaag gatgagtaag tatgggccca gccagatgag gagcaccgtg
     7861 gtggaagcag agagcggggt gaggccccta gtgagggggg ctgcctgtgc ttcggggcct
     7921 tacactgctc tttggggtgc agccaaccct tccctgcgcc atgggagcct ccgtacccac
     7981 cttccctgtg cagtcactcc cccgcagtct cctgctcaga ccctcctcac cccccaggtt
     8041 caagcagctt ctcctgaccc aggcagacaa gttctctcca gctgaggtga ggctgcccag
     8101 ccccttcaat actcatcccc agcaccttct ctgggccttc acccatgacc cagagcccag
     8161 taccagtgag gcagttgctg gaagggtgag ccgagggccc ttctggagga ggtgccatct
     8221 ctgttgagac ctagagggta aagatgtgga gtcagaaaag agggcagggt gcgccaggca
     8281 gggagactgt gcacagacct ggggggaagt ggatagggag aggtttcgta cactcggggt
     8341 gggcctgtgc ctgtggctgg aggggcgtcc tttgcc
//
