LOCUS       sequence-231775-        7157 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing iBlueberry and mCherry in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7157)
  AUTHORS   Yu D, Dong Z, Gustafson WC, Ruiz-Gonzalez R, Signor L, Marzocca F, 
            Borel F, Klassen MP, Makhijani K, Royant A, Jan YN, Weiss WA, Guo S,
            Shu X
  TITLE     Rational design of a monomeric and photostable far-red fluorescent 
            protein for fluorescence imaging in vivo.
  JOURNAL   Protein Sci. 2016 Feb;25(2):308-15. doi: 10.1002/pro.2843. Epub 2015
            Dec 31.
  PUBMED    26549191
REFERENCE   2  (bases 1 to 7157)
  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: "10.1002/pro.2843"; 
            journalName: "Protein Sci"; date: "2016-02"; volume: "25"; issue: 
            "2"; pages: "308-15"
COMMENT     Sequence Label: pcDNA3.1_LifeAct-iBlueberry
FEATURES             Location/Qualifiers
     source          1..7157
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     865..884
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        865..883
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      919..928
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             925..975
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     CDS             982..1941
                     /codon_start=1
                     /product="naturally monomeric infrared fluorescent protein 
                     derived from a truncated bacteriophytochrome (Yu et al., 
                     2015)"
                     /label=mIFP
                     /translation="MSVPLTTSAFGHAFLANCEREQIHLAGSIQPHGILLAVKEPDNVV
                     IQASINAAEFLNTNSVVGRPLRDLGGDLPLQILPHLNGPLHLAPMTLRCTVGSPPRRVD
                     CTIHRPSNGGLIVELEPATKTTNIAPALDGAFHRITSSSSLMGLCDETATIIREITGYD
                     RVMVVRFDEEGNGQILSERRRADLEAFLGNRYPASTIPQIARRLYEHNRVRLLVDVNYT
                     PVPLQPRISPLNGRDLDMSLSCLRSMSPCHQKYMQDMGVGATLVCSLMVSGRLWGLIAC
                     HHYEPRFVPFHIRAAGEALAETCAIRIATLESFAQSQSK"
     CDS             1948..2001
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2008..2718
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2153..2171)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2421..2440
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2750..2767)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2756..2980
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3026..3454
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3113..3132)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3323..3344
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3463..3483)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3468..3797
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3648..3783
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3710..3729
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3864..4658
                     /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(3918..3937)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4528..4547
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4832..4953
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4869..4888)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4923..4942
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5002..5018)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5002..5018)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5015..5037)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5026..5042
                     /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(5050..5056,5057..5074,5075..5080))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5095..5116
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5233..5250)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5404..5992)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5484..5503)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6163..6954,6955..7023))
                     /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     6786..6805
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7024..7128)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      841 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta
      901 gcgtttaaac ttaagcttgc caccatgggc gtggccgact tgatcaagaa gttcgagtcc
      961 atctccaagg aggagggcag catgtcggta ccgctgacta cctcagcatt cggccacgcg
     1021 tttctggcta actgtgaacg cgagcagatc cacctggcgg gctccattca gccgcacggt
     1081 atcctgctgg ctgtgaaaga gccggacaac gtggtgatcc aggcttctat taacgctgcg
     1141 gagttcctga acaccaactc tgttgttggc cgtccgctgc gtgacctggg cggcgatctg
     1201 cctttgcaga tcctgccgca cctgaacggc ccgctgcacc tggctccgat gaccctgcgt
     1261 tgtaccgtgg gttctccgcc gcgtcgtgtg gactgtacca ttcatcgtcc gtctaacggc
     1321 ggcctgatcg tagaactgga accagcaacc aagaccacta acattgcgcc ggctctggac
     1381 ggtgcgtttc atcgtatcac ttcttcatcc tccctgatgg gcctgtgtga cgaaaccgcg
     1441 actattatcc gtgagattac tggctacgac cgtgtgatgg tagtacgttt cgatgaagag
     1501 ggtaatggcc aaattctgtc cgaacgtcgt cgtgcggacc tggaagcgtt cctgggtaac
     1561 cgctacccgg cgtctactat tccgcagatc gctcgtcgcc tgtacgaaca taaccgtgtt
     1621 cgcctgctgg tagatgtgaa ctatactccg gttccgctac agccgcgcat cagcccgctg
     1681 aacggtcgtg atctggatat gtccctgtct tgcctgcgct ctatgtcccc gtgccaccag
     1741 aaatacatgc aggacatggg cgttggcgcg accctggttt gctctctgat ggtgtctggt
     1801 cgtctgtggg gtctgatcgc ttgccaccac tacgaaccgc gcttcgttcc gttccacatt
     1861 cgcgctgctg gcgaagcgct ggcggaaact tgtgcgatcc gcatcgcgac gctggagagc
     1921 tttgcacagt ctcagtccaa aggatccgag ggcagaggaa gtctgctaac atgcggtgac
     1981 gtcgaggaga atcctggccc agaattcatg gtgagcaagg gcgaggagga taacatggcc
     2041 atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag
     2101 ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg
     2161 aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg
     2221 tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt gaagctgtcc
     2281 ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc
     2341 gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa gctgcgcggc
     2401 accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc
     2461 tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca gaggctgaag
     2521 ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc caagaagccc
     2581 gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc ccacaacgag
     2641 gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg
     2701 gacgagctgt acaagtaatc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg
     2761 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     2821 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     2881 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     2941 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     3001 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3061 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3121 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3181 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3241 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3301 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     3361 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     3421 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     3481 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     3541 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3601 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     3661 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     3721 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     3781 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag
     3841 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     3901 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     3961 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     4021 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     4081 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     4141 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     4201 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     4261 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     4321 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     4381 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     4441 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     4501 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     4561 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     4621 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     4681 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     4741 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     4801 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     4861 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     4921 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     4981 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     5041 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     5101 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     5161 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     5221 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     5281 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     5341 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     5401 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5461 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     5521 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5581 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5641 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5701 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5761 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5821 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     5881 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5941 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     6001 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     6061 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     6121 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     6181 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     6241 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     6301 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     6361 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     6421 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     6481 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     6541 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     6601 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     6661 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     6721 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     6781 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     6841 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     6901 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6961 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     7021 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     7081 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     7141 aaaagtgcca cctgacg
//
