LOCUS       sequence-414808-        7493 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tagged MDFI for western blotting and imaging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7493)
  AUTHORS   Zhou Z, Ma X, Lin Y, Cheng D, Bavi N, Secker GA, Li JV, Janbandhu V,
            Sutton DL, Scott HS, Yao M, Harvey RP, Harvey NL, Corry B, Zhang Y, 
            Cox CD
  TITLE     MyoD-family inhibitor proteins act as auxiliary subunits of Piezo 
            channels.
  JOURNAL   Science. 2023 Aug 18;381(6659):799-804. doi: 
            10.1126/science.adh8190. Epub 2023 Aug 17.
  PUBMED    37590348
REFERENCE   2  (bases 1 to 7493)
  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: 
            "10.1126/science.adh8190"; journalName: "Science"; date: 
            "2023-08-18- 18"; volume: "381"; issue: "6659"; pages: "799-804"
COMMENT     Sequence Label: HA-MDFI
FEATURES             Location/Qualifiers
     source          1..7493
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             926..952
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    join(1732..2306,2307..2309,2310..2318)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1899..1916)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2126..2145
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2319..2321,2322..2324,2325..3038)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2319..2321,2322..2324,2325..3035)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2364..2385)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2625..2644)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2972..2993
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3084..3101)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3090..3314
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3360..3788
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3447..3466)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3657..3678
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3797..3817)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3802..4131
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3982..4117
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4044..4063
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4198..4992
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4252..4271)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4862..4881
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5166..5287
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5203..5222)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5257..5276
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5336..5352)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5336..5352)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5349..5371)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5360..5376
                     /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(5384..5390,5391..5408,5409..5414))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5429..5450
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5567..5584)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5738..6326)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5818..5837)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6497..7288,7289..7357))
                     /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     7120..7139
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7358..7462)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttgcca ccatgtaccc atacgatgtt ccagattacg ctatgtacca
      961 ggtgagcggc cagcgcccct ctggctgcga cgcgccctat ggagccccca gcgcagcccc
     1021 gggcccagcc cagaccctat ccctccttcc tgggctggag gtagtaacag gatccactca
     1081 ccctgcggag gcagcaccag aggagggctc cctggaggag gcggcaaccc ccatgcccca
     1141 aggcaatggc cctggcatcc cccagggcct ggacagcact gacctcgacg tccccacaga
     1201 agctgtgaca tgccagcctc aggggaaccc cttgggctgc accccacttc tgccgaatga
     1261 ctctggccac ccctcagagc tgggcggcac cagacgggcg gggaatggtg ccctgggtgg
     1321 ccccaaggcc caccggaagt tgcagacaca cccatctctc gccagccagg gcagcaagaa
     1381 gagtaagagc agcagcaaat ccaccacctc ccagatcccc ctccaggcac aggaagactg
     1441 ctgtgtccac tgcatcctgt cctgcctgtt ctgcgagttc ctgacgctgt gcaacatcgt
     1501 cctggactgc gccacctgtg gctcctgcag ctcggaggac tcgtgcctct gctgctgctg
     1561 ctgtggctct ggcgagtgtg ccgactgcga cctgccctgc gacctggact gcggcatcct
     1621 ggatgcctgc tgcgagtccg cggactgcct ggagatctgc atggagtgct gtgggctctg
     1681 cttctcctcc tgagaattct gcagtcgacg gtaccgcggg cccgggatcc gcccctctcc
     1741 ctcccccccc cctaacgtta ctggccgaag ccgcttggaa taaggccggt gtgcgtttgt
     1801 ctatatgtta ttttccacca tattgccgtc ttttggcaat gtgagggccc ggaaacctgg
     1861 ccctgtcttc ttgacgagca ttcctagggg tctttcccct ctcgccaaag gaatgcaagg
     1921 tctgttgaat gtcgtgaagg aagcagttcc tctggaagct tcttgaagac aaacaacgtc
     1981 tgtagcgacc ctttgcaggc agcggaaccc cccacctggc gacaggtgcc tctgcggcca
     2041 aaagccacgt gtataagata cacctgcaaa ggcggcacaa ccccagtgcc acgttgtgag
     2101 ttggatagtt gtggaaagag tcaaatggct ctcctcaagc gtattcaaca aggggctgaa
     2161 ggatgcccag aaggtacccc attgtatggg atctgatctg gggcctcggt acacatgctt
     2221 tacatgtgtt tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg gggacgtggt
     2281 tttcctttga aaaacacgat gataatatgg ccacaaccat ggtgagcaag ggcgaggagc
     2341 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     2401 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     2461 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     2521 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     2581 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     2641 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     2701 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
     2761 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     2821 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     2881 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     2941 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     3001 ccgggatcac tctcggcatg gacgagctgt acaagtaaag cggccgctcg agtctagagg
     3061 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     3121 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     3181 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     3241 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     3301 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     3361 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     3421 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     3481 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     3541 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     3601 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     3661 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     3721 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     3781 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3841 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3901 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     3961 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4021 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     4081 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     4141 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     4201 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     4261 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4321 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     4381 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4441 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4501 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     4561 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4621 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4681 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     4741 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4801 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4861 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4921 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4981 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     5041 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     5101 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     5161 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     5221 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     5281 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     5341 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     5401 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     5461 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     5521 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     5581 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     5641 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     5701 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     5761 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     5821 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5881 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5941 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     6001 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     6061 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     6121 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     6181 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     6241 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     6301 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     6361 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     6421 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     6481 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6541 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6601 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6661 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6721 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6781 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6841 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6901 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6961 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     7021 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     7081 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     7141 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     7201 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     7261 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     7321 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     7381 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     7441 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
