LOCUS       sequence-192387-        7455 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To clone the 3 ORFs for Gal4 dependent generation of ifgMosaics.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7455)
  AUTHORS   Pontes-Quero S, Heredia L, Casquero-Garcia V, Fernandez-Chacon M, 
            Luo W, Hermoso A, Bansal M, Garcia-Gonzalez I, Sanchez-Munoz MS, 
            Perea JR, Galiana-Simal A, Rodriguez-Arabaolaza I, Del Olmo-Cabrera 
            S, Rocha SF, Criado-Rodriguez LM, Giovinazzo G, Benedito R
  TITLE     Dual ifgMosaic: A Versatile Method for Multispectral and 
            Combinatorial Mosaic Gene-Function Analysis.
  JOURNAL   Cell. 2017 Aug 10;170(4):800-814.e18. doi: 
            10.1016/j.cell.2017.07.031.
  PUBMED    28802047
REFERENCE   2  (bases 1 to 7455)
  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.cell.2017.07"; journalName: "Cell"; date: "2017-08-10- 
            10"; volume: "170"; issue: "4"; pages: "800-814.e18"
COMMENT     Sequence Label: 4xnr UAS ifgMosaic Donor vector (IG230)
FEATURES             Location/Qualifiers
     source          1..7455
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     protein_bind    complement(606..639)
                     /label=loxN
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     protein_bind    complement(647..680)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(689..722)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      736..745
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             781..801
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             802..1506
                     /codon_start=1
                     /product="Phialidium yellow fluorescent proteim"
                     /label=PhiYFP
                     /note="mammalian codon-optimized"
                     /translation="MSSGALLFHGKIPYVVEMEGNVDGHTFSIRGKGYGDASVGKVDAQ
                     FICTTGDVPVPWSTLVTTLTAGAQCFAKYGPELKDFYKSCMPDGYVQERTITFEGDGNF
                     KTRAEVTFENGSVYNRVKLNGQGFKKDGHVLGKNLEFNFTPHCLYIWGDQANHGLKSAF
                     KICHEITGSKGDFIVADHTQMNTPIGGGPVHVPEYHHMSYHVKLSKDVTDHRDNMSLKE
                     TVRAVDCRKTYL"
     polyA_signal    1689..1810
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1726..1745)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1780..1799
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1874..1907
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     promoter        1971..2470
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2006..2029)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2403..2422
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        2482..2529
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2548..3351
                     /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(2611..2630)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3221..3240
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3383..3400)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3389..3613
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3671..3704
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(5234..5250)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5234..5250)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5247..5269)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5258..5274
                     /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(5282..5288,5289..5306,5307..5312))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5327..5348
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5465..5482)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5636..6224)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5716..5735)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6395..7186,7187..7255))
                     /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     7018..7037
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7256..7360)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7428..7446
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgccgg cggtggcttc taatccgtga gtcctagcgg
      421 gtgacagccc tccgtcttca caggcggagg agagtcttcc gtagggttcc tcggagtact
      481 gtcctccgac gcgtgcaagg gtcgactcta gagggtatat aatggatccc atcgcgtctc
      541 agcctcactt tgagctcctc cacacgaatt cgccactttg taagcttggc aaagaattcc
      601 tcgagataac ttcgtatagt ataccttata cgaagttatc gagctcataa cttcgtatag
      661 gatactttat acgaagttat aactcgagat aacttcgtat agcatacatt atacgaagtt
      721 ataccggcag atctcgccac catggtgagt gtgattacag aggctagctt caccatggct
      781 cctaagaaga agaggaaggt gatgagcagc ggcgccctgc tgttccacgg caagatcccc
      841 tacgtggtgg agatggaggg caatgtggat ggccacacct tcagcatccg cggcaagggc
      901 tacggcgatg ccagcgtggg caaggtggat gcccagttca tctgcaccac aggcgatgtg
      961 cccgtgccct ggagcaccct ggtgaccacc ctgaccgcag gcgcccagtg cttcgccaag
     1021 tacggccccg agctgaagga tttctacaag agctgcatgc ccgatggcta cgtgcaggag
     1081 cgcaccatca ccttcgaggg tgatggcaat ttcaagaccc gcgccgaggt gaccttcgag
     1141 aatggcagcg tgtacaatcg cgtgaagctg aatggccagg gcttcaagaa ggatggccac
     1201 gtgctgggca agaatctgga gttcaatttc accccccact gcctgtacat ctggggcgat
     1261 caggccaatc acggcctgaa gagcgccttc aagatctgcc acgagatcac cggcagcaag
     1321 ggcgatttca tcgtggccga tcacacccag atgaataccc ccatcggcgg cggccccgtg
     1381 cacgtgcccg agtaccacca catgagctac cacgtgaagc tgagcaagga tgtgaccgat
     1441 caccgcgata atatgagcct gaaggagacc gtgcgcgccg tggattgccg caagacctac
     1501 ctgtgacgct taattagctg agcttggact cctgttgata gatccagtaa tgacctcaga
     1561 actccatctg gccgcgactc tagatcataa tcagccatac cacatttgta gaggttttac
     1621 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     1681 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     1741 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     1801 atgtatctta aggcgaagct tcagccctat cattcggcgc gccgcggccg cagcataggg
     1861 atccccgaat tccgaagttc ctattctcta gaaagtatag gaacttcagg tctgaagagg
     1921 agtttacgtc cagccaagct agcttggctg caggtcgtcg aaattctacc gggtagggga
     1981 ggcgcttttc ccaaggcagt ctggagcatg cgctttagca gccccgctgg gcacttggcg
     2041 ctacacaagt ggcctctggc ctcgcacaca ttccacatcc accggtaggc gccaaccggc
     2101 tccgttcttt ggtggcccct tcgcgccacc ttctactcct cccctagtca ggaagttccc
     2161 ccccgccccg cagctcgcgt cgtgcaggac gtgacaaatg gaagtagcac gtctcactag
     2221 tctcgtgcag atggacagca ccgctgagca atggaagcgg gtaggccttt ggggcagcgg
     2281 ccaatagcag ctttgctcct tcgctttctg ggctcagagg ctgggaaggg gtgggtccgg
     2341 gggcgggctc aggggcgggc tcaggggcgg ggcgggcgcc cgaaggtcct ccggaggccc
     2401 ggcattctgc acgcttcaaa agcgcacgtc tgccgcgctg ttctcctctt cctcatctcc
     2461 gggcctttcg acctgcagcc tgttgacaat taatcatcgg catagtatat cggcatagta
     2521 taatacgaca aggtgaggaa ctaaaccatg ggatcggcca ttgaacaaga tggattgcac
     2581 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     2641 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     2701 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     2761 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     2821 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     2881 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     2941 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     3001 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     3061 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg atgatctcgt cgtgacccat
     3121 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     3181 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     3241 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     3301 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg aggggatcaa
     3361 ttctctagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     3421 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     3481 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     3541 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     3601 gtgggctcta tggcttctga ggcggaaaga accagctggg gctcgactag agcttgcgga
     3661 acccttcgtt gaagttccta ttctctagaa agtataggaa cttcagtcga cgtgctccaa
     3721 ggagagtttg ctggcaagat cttcggggtc gatgctaaca cagggaatgt gcttgcctat
     3781 gagaggctag accgggagaa agtctctgag tacttcctta ctgccctcat tgtggacaag
     3841 aacaccaaca aaaacctgga acaaccttcc agcttcactg tcaaggtgca tgatataaat
     3901 gacaattggc ctgtgttttc gcaccaggta ttcaacgcat ctgtgccaga gatgtcagct
     3961 ataggtatgc acctagtctt ccctctctcc tcccttcctc aacaccagcc tgaagacacc
     4021 tctcacatca gccacctcaa tgcttagggt caggctccag taagctcaag gaagtagttg
     4081 taatgctatc ttgtgccttt acatgatttc aaaatgtgtc cctaaaggct aaactctgac
     4141 ctagccccaa aataaaccct gaccctaatc atagatgggt tttaatcttc acaatgtgct
     4201 gaacactgac tctcattata gcctgggcct tgatcctgac cgcagagtgt gctgtcaccc
     4261 tagatatggg cttaggaggt actagcctgt cttcatcctg cttacagaag gtccaggcaa
     4321 agagtgtgtc tctctctact cttctgtctg ggatctgtgc tcaaggccat ggagccagac
     4381 cttcttgagg aaacccagaa ctccaaagga tgagaaatgg aatctcaacc cctctgggaa
     4441 ctttgcaaca cccttcctag tcccgcattc atactttcag aaaatgaaaa tctacccaga
     4501 cgtcacctat gtttctctct caaacccaac cccttccctt cttcatttct cccacaaaca
     4561 cacacaccac ctgttctctc tgcctgcttc atgactgctc cggctggcac tttgtcctca
     4621 tgatggcccc tctagcacaa ggcagccttc attccctcat cttgggcagt aggaacagca
     4681 gaacccccat ctactcccac caggaaatgg ggtgagggcc agtcatgagc ggacatacca
     4741 gaattctctc tctccacagg gacctctgtc atccgtgtga cagcagtgga tgcagatgac
     4801 cccactgtgg ccggccacgc cactgtcttg taccaaatcg tgaaaggaaa tgagtatttc
     4861 agcattgata attctggtct gtatgactaa gccttttggg cgggtaccca tttggaggga
     4921 tggaggaggg acgtgaatat tgcaggtacc agtgtataga gaacattaac ctgataggtg
     4981 tcaccagcca ctgatccaga gaaggaatgc ataatacaca taaagacatg ttggagtggg
     5041 ttaggggaga aggaagcaga caagcgccca aagcaggacc tcaagtccag aaagaagatc
     5101 ggggcctttt cacccaagca tagaccagga ctcaacagaa catgctggtt ggaaagtctt
     5161 ttgtacacat tcagagacat ctgatttgaa cccagaacta ctgctcagcc cttaattaag
     5221 aattcgtaat catgtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5281 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5341 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     5401 tgcattaatg aatcgaccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     5461 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5521 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5581 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5641 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5701 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5761 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5821 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5881 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5941 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     6001 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     6061 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     6121 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6181 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6241 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6301 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6361 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6421 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6481 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6541 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6601 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6661 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6721 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6781 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6841 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6901 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6961 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     7021 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     7081 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     7141 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7201 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7261 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7321 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     7381 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     7441 cgaggccctt tcgtc
//
