LOCUS       sequence-196886-        6703 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bicistronic expression of mRFP-Utrophin and Neo resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6703)
  TITLE     Piel Lab Plasmids
REFERENCE   2  (bases 1 to 6703)
  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"
COMMENT     Sequence Label: pIRESneo3+mRFP-Utr_CH
FEATURES             Location/Qualifiers
     source          1..6703
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             702..1376
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             702..1376
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(832..850)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1100..1119
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1294..1317
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     intron          2255..2484
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2467..2486
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    2658..3120
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2714..2731)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2941..2960
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3156..3959
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3219..3238)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3829..3848
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4089..4210
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4126..4145)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4312..4328)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4312..4328)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4325..4347)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4336..4352
                     /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(4360..4366,4367..4384,4385..4390))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4405..4426
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4543..4560)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4714..5302)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4794..4813)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5473..6264,6265..6333))
                     /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     6096..6115
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6334..6438)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6513..6532)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccgagc tcggatcgat atctgcggcc tagctagcac catggcctcc tccgaggacg
      721 tcatcaagga gttcatgcgc ttcaaggtgc gcatggaggg ctccgtgaac ggccacgagt
      781 tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga
      841 aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtcccct cagttccagt
      901 acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg aagctgtcct
      961 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg
     1021 tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag ctgcgcggca
     1081 ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg gaggcctcca
     1141 ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagatg aggctgaagc
     1201 tgaaggacgg cggccactac gacgccgagg tcaagaccac ctacatggcc aagaagcccg
     1261 tgcagctgcc cggcgcctac aagaccgaca tcaagctgga catcacctcc cacaacgagg
     1321 actacaccat cgtggaacag tacgagcgcg ccgagggccg ccactccacc ggcgcctccg
     1381 gaaccatggc caagtatgga gaacatgaag ccagtcctga caatgggcag aacgaattca
     1441 gtgatatcat taagtccaga tctgatgaac acaatgacgt acagaagaaa acctttacca
     1501 aatggataaa tgctcgattt tcaaagagtg ggaaaccacc catcaatgat atgttcacag
     1561 acctcaaaga tggaaggaag ctattggatc ttctagaagg cctcacagga acatcactgc
     1621 caaaggaacg tggttccaca agggtacatg ccttaaataa cgtcaacaga gtgctgcagg
     1681 ttttacatca gaacaatgtg gaattagtga atataggggg aaccgacatt gtggatggaa
     1741 atcacaaact gactttgggg ttactttgga gcatcatttt gcactggcag gtgaaagatg
     1801 tcatgaagga tgtcatgtcg gacctgcagc agacgaacag tgagaagatc ctgctcagct
     1861 gggtgcgtca gaccaccagg ccctacagcc aagtcaacgt cctcaacttc accaccagct
     1921 ggacagatgg actcgccttt aatgctgtcc tccaccgaca taaacctgat ctcttcagct
     1981 gggataaagt tgtcaaaatg tcaccaattg agagacttga acatgccttc agcaaggctc
     2041 aaacttattt gggaattgaa aagctgttag atcctgaaga tgttgccgtt cggcttcctg
     2101 acaagaaatc cataattatg tatttaacat ctttgtttga ggtgctacct cagcaagtca
     2161 ccatagacta attcgaattc ggatccgcgg ccgcatagat aactgatcca gtgtgctgga
     2221 attaattcgc tgtctgcgag ggccagctgt tggggtgagt actccctctc aaaagcgggc
     2281 atgacttctg cgctaagatt gtcagtttcc aaaaacgagg aggatttgat attcacctgg
     2341 cccgcggtga tgcctttgag ggtggccgcg tccatctggt cagaaaagac aatctttttg
     2401 ttgtcaagct tgaggtgtgg caggcttgag atctggccat acacttgagt gacaatgaca
     2461 tccactttgc ctttctctcc acaggtgtcc actcccaggt ccaactgcag gtcgagcatg
     2521 catctagggc ggccaattcc gcccctctcc cccccccccc ccccccctaa cgttactggc
     2581 cgaagccgct tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg
     2641 ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct
     2701 aggggtcttt cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca
     2761 gttcctctgg aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg
     2821 aaccccccac ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct
     2881 gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa
     2941 tggctctcct caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt
     3001 atgggatctg atctggggcc tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa
     3061 aaacgtctag gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgatgataa
     3121 gcttgccaca acccgggata attcctgcag ccaatatggg atcggccatt gaacaagatg
     3181 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     3241 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     3301 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     3361 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     3421 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     3481 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     3541 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     3601 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     3661 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgat gatctcgtcg
     3721 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     3781 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     3841 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     3901 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     3961 gggatcaatt ctctagataa ctgatcataa tcagccatac cacatttgta gaggttttac
     4021 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     4081 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     4141 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     4201 atgtatctta acgcgtcgag tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4261 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     4321 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4381 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     4441 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4501 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4561 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4621 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4681 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4741 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4801 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4861 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4921 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4981 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5041 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5101 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5161 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5221 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5281 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5341 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5401 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5461 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5521 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     5581 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5641 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5701 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5761 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5821 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5881 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5941 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6001 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6061 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6121 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6181 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6241 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6301 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6361 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6421 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg
     6481 agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     6541 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
     6601 taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg
     6661 cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gtt
//
