LOCUS       sequence-456542-        5917 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Overexpression of human MYOD1 ORF with a 3xHA C-terminal tag and an 
            IRES-hrGFP reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5917)
  TITLE     Alexander lab Muscle Factors
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5917)
  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: 
            "Unpublished"
COMMENT     Sequence Label: hMYOD1-3xHA-IRES-hrGFP (#343)
FEATURES             Location/Qualifiers
     source          1..5917
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             309..389
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     misc_feature    join(425..996,997..999,1000..1008)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(592..609)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     819..838
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             1009..1728
                     /codon_start=1
                     /product="humanized Renilla green fluorescent protein"
                     /label=hrGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKQILKNTGLQEIMSFKVNLEGVVNNHVFTMEGCGKGNILFGN
                     QLVQIRVTKGAPLPFAFDILSPAFQYGNRTFTKYPEDISDFFIQSFPAGFVYERTLRYE
                     DGGLVEIRSDINLIEEMFVYRVEYKGRNFPNDGPVMKKTITGLQPSFEVVYMNDGVLVG
                     QVILVYRLNSGKFYSCHMRTLMKSKGVVKDFPEYHFIQHRLEKTYVEDGGFVEQHETAI
                     AQLTSLGKPLGSLHEWV"
     primer_bind     complement(1758..1777)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1759..1777)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2051..2172
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2088..2107)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2142..2161
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2179..2634)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2316..2337)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2528..2547
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2661..2765
                     /gene="bla"
                     /label=AmpR promoter
     protein_bind    2780..2813
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(2855..3646,3647..3715))
                     /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     3478..3497
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3885..4473
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4374..4393
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4648..4951
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4952..5155
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5106..5126
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        5201..5219
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      5254..5263
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 cccggcggcg gaactgctac gaaggcgcct actacaacga ggcgcccagc gaacccaggc
       61 ccgggaagag tgcggcggtg tcgagcctag actgcctgtc cagcatcgtg gagcgcatct
      121 ccaccgagag ccctgcggcg cccgccctcc tgctggcgga cgtgccttct gagtcgcctc
      181 cgcgcaggca agaggctgcc gcccccagcg agggagagag cagcggcgac cccacccagt
      241 caccggacgc cgccccgcag tgccctgcgg gtgcgaaccc caacccgata taccaggtgc
      301 tcctcgagta cccatatgac gttccagact acgcgtatcc gtacgacgtt ccggattacg
      361 cttaccctta cgatgtacct gactacgctt aatagcaatt cctcgacgac tgcatagggt
      421 tacccccctc tccctccccc ccccctaacg ttactggccg aagccgcttg gaataaggcc
      481 ggtgtgcgtt tgtctatatg ttattttcca ccatattgcc gtcttttggc aatgtgaggg
      541 cccggaaacc tggccctgtc ttcttgacga gcattcctag gggtctttcc cctctcgcca
      601 aaggaatgca aggtctgttg aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa
      661 gacaaacaac gtctgtagcg accctttgca ggcagcggaa ccccccacct ggcgacaggt
      721 gcctctgcgg ccaaaagcca cgtgtataag atacacctgc aaaggcggca caaccccagt
      781 gccacgttgt gagttggata gttgtggaaa gagtcaaatg gctctcctca agcgtattca
      841 acaaggggct gaaggatgcc cagaaggtac cccattgtat gggatctgat ctggggcctc
      901 ggtgcacatg ctttacatgt gtttagtcga ggttaaaaaa cgtctaggcc ccccgaacca
      961 cggggacgtg gttttccttt gaaaaacacg atgataatgg ccacaaccat ggtgagcaag
     1021 cagatcctga agaacaccgg cctgcaggag atcatgagct tcaaggtgaa cctggagggc
     1081 gtggtgaaca accacgtgtt caccatggag ggctgcggca agggcaacat cctgttcggc
     1141 aaccagctgg tgcagatccg cgtgaccaag ggcgcccccc tgcccttcgc cttcgacatc
     1201 ctgagccccg ccttccagta cggcaaccgc accttcacca agtaccccga ggacatcagc
     1261 gacttcttca tccagagctt ccccgccggc ttcgtgtacg agcgcaccct gcgctacgag
     1321 gacggcggcc tggtggagat ccgcagcgac atcaacctga tcgaggagat gttcgtgtac
     1381 cgcgtggagt acaagggccg caacttcccc aacgacggcc ccgtgatgaa gaagaccatc
     1441 accggcctgc agcccagctt cgaggtggtg tacatgaacg acggcgtgct ggtgggccag
     1501 gtgatcctgg tgtaccgcct gaacagcggc aagttctaca gctgccacat gcgcaccctg
     1561 atgaagagca agggcgtggt gaaggacttc cccgagtacc acttcatcca gcaccgcctg
     1621 gagaagacct acgtggagga cggcggcttc gtggagcagc acgagaccgc catcgcccag
     1681 ctgaccagcc tgggcaagcc cctgggcagc ctgcacgagt gggtgtaata gggtaccagg
     1741 taagtgtacc caattcgccc tatagtgagt cgtattacaa ttcactcgat cgcccttccc
     1801 aacagttgcg cagcctgaat ggcgaatgga gatccaattt ttaagtgtat aatgtgttaa
     1861 actactgatt ctaattgttt gtgtatttta gattcacagt cccaaggctc atttcaggcc
     1921 cctcagtcct cacagtctgt tcatgatcat aatcagccat accacatttg tagaggtttt
     1981 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat
     2041 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     2101 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     2161 caatgtatct taacgcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     2221 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     2281 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     2341 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     2401 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     2461 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     2521 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     2581 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg
     2641 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     2701 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     2761 agaatcctga ggccgggcca taacttcgta taatgtatgc tatacgaagt tatccatggg
     2821 ccccccctcg acatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     2881 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     2941 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     3001 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     3061 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     3121 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     3181 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     3241 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     3301 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     3361 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     3421 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     3481 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     3541 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     3601 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     3661 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     3721 tcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     3781 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     3841 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     3901 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     3961 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     4021 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     4081 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     4141 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     4201 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     4261 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     4321 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     4381 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     4441 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     4501 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     4561 gtggataacc gtattaccgc catgcattag ttattaatag taatcaatta cggggtcatt
     4621 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     4681 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     4741 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     4801 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     4861 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     4921 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     4981 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     5041 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     5101 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctggttt
     5161 agtgaaccgt cagatccgct agcgattacg ccaagctcga aattaaccct cactaaaggg
     5221 aacaaaagct ggagctccac cgcggtggcg gccgccacca tggagctact gtcgccaccg
     5281 ctccgcgacg tagacctgac ggcccccgac ggctctctct gctcctttgc cacaacggac
     5341 gacttctatg acgacccgtg tttcgactcc ccggacctgc gcttcttcga agacctggac
     5401 ccgcgcctga tgcacgtggg cgcgctcctg aaacccgaag agcactcgca cttccccgcg
     5461 gcggtgcacc cggccccggg cgcacgtgag gacgagcatg tgcgcgcgcc cagcgggcac
     5521 caccaggcgg gccgctgcct actgtgggcc tgcaaggcgt gcaagcgcaa gaccaccaac
     5581 gccgaccgcc gcaaggccgc caccatgcgc gagcggcgcc gcctgagcaa agtaaatgag
     5641 gcctttgaga cactcaagcg ctgcacgtcg agcaatccaa accagcggtt gcccaaggtg
     5701 gagatcctgc gcaacgccat ccgctatatc gagggcctgc aggctctgct gcgcgaccag
     5761 gacgccgcgc cccctggcgc cgcagccgcc ttctatgcgc cgggcccgct gcccccgggc
     5821 cgcggcggcg agcactacag cggcgactcc gacgcgtcca gcccgcgctc caactgctcc
     5881 gacggcatga tggactacag cggccccccg agcggcg
//
