LOCUS       sequence-360261-        5157 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for PUF9R-iRFP670.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5157)
  AUTHORS   Clow PA, Du M, Jillette N, Taghbalout A, Zhu JJ, Cheng AW
  TITLE     CRISPR-mediated multiplexed live cell imaging of nonrepetitive 
            genomic loci with one guide RNA per locus.
  JOURNAL   Nat Commun. 2022 Apr 6;13(1):1871. doi: 10.1038/s41467-022-29343-z.
  PUBMED    35387989
REFERENCE   2  (bases 1 to 5157)
  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: "Nat 
            Commun."; date: "2022-04-6"; volume: "13(1):1871. doi"; pages: " 
            10.1038/s41467-022-29343-z"
COMMENT     Sequence Label: pAC11002-pmax-PUF9R_iRFP670
FEATURES             Location/Qualifiers
     source          1..5157
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          813..945
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     protein_bind    960..984
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1002..1011
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1023..1043
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2235..3170
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 670 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP670
                     /note="derived from Rhodopseudomonas palustris RpBphP6"
                     /translation="MARKVDLTSCDREPIHIPGSIQPCGCLLACDAQAVRITRITENAG
                     AFFGRETPRVGELLADYFGETEAHALRNALAQSSDPKRPALIFGWRDGLTGRTFDISLH
                     RHDGTSIIEFEPAAAEQADNPLRLTRQIIARTKELKSLEEMAARVPRYLQAMLGYHRVM
                     LYRFADDGSGMVIGEAKRSDLESFLGQHFPASLVPQQARLLYLKNAIRVVSDSRGISSR
                     IVPEHDASGAALDLSFAHLRSISPCHLEFLRNMGVSASMSLSIIIDGTLWGLIICHHYE
                     PRAVPMAQRVAAEMFADFLSLHFTAAHHQR"
     protein_bind    complement(3186..3210)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3263..3304
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3330..3349)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    3359..3440
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     3384..3403
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3638..4226)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3718..3737)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4307..5101)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(4307..5101)
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4418..4437)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5028..5047
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagg
      721 tcgtttagtg aaccgtcaga tcactagtag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctc gactgatcac aggtaagtat caaggttaca agacaggttt
      841 aaggaggcca atagaaactg ggcttgtcga gacagagaag attcttgcgt ttctgatagg
      901 cacctattgg tcttactgac atccactttg cctttctctc cacagggaaa aaacaattga
      961 caagtttgta caaaaaagca ggctccgaat tcaccggtgc cgccaccatg gggatcctcc
     1021 cccccaagaa aaagaggaag gtatctagag gccgcagccg ccttttggaa gattttcgaa
     1081 acaaccggta ccccaattta caactgcggg agattgctgg acatataatg gaattttccc
     1141 aagaccagca tgggtccaga ttcattcagc tgaaactgga gcgtgccaca ccagctgagc
     1201 gccagcttgt cttcaatgaa atcctccagg ctgcctacca actcatggtg gatgtgtttg
     1261 gtaattacgt cattcagaag ttctttgaat ttggcagtct tgaacagaag ctggctttgg
     1321 cagaacggat tcgaggccac gtcctgtcat tggcactaca gatgtatggc tgccgtgtta
     1381 tccagaaagc tcttgagttt attccttcag accagcagaa tgagatggtt cgggaactcc
     1441 accaacacac ggagcaactg gtccaggatc aatacggcaa ctacgtgatt cagcacgtgt
     1501 tggagcacgg taggccagag gacaagagta aaattgtcgc cgagatccgg ggcaatgttc
     1561 tggtcctctc tcagcacaaa ttcgcctcca atgttgtgga aaagtgtgtg acccacgcat
     1621 ccagaactga acgagctgtg ctcatcgacg aggtttgtac gatgaatgac ggtcctcact
     1681 ctgcacttta tacaatgatg aaggaccagt acgccaatta tgttgtgcag aaaatgatcg
     1741 acgtagcaga acccggacag cgcaaaatcg tgatgcacga gcttcaccag cacacagagc
     1801 agctggtaca ggatcaatat ggaaattatg taatccaaca tgtactggag cacggtcgtc
     1861 ctgaggataa aagcaaaatt gtagcagaaa tccgaggcaa tgtacttgta ttgagtcagc
     1921 acaaatttgc aagcaatgtt gtggagaagt gtgttactca cgcctcacgt acggagcgcg
     1981 ctgtgctcat cgacgaggtg tgcaccatga acgacggtcc ccacagtgcc ttatacacca
     2041 tgatgaagga ccagtatgcc aactacgtgg tccagaagat gattgacgtg gcggagccag
     2101 gccagcggaa gatcgtcatg cataagatcc ggccccacat cgcaactctt cgtaagtaca
     2161 cctatggcaa gcacattctg gccaagctgg agaagtacta catgaagaac ggtgttgact
     2221 taggggggcc ggccatggcg cgtaaggtcg atctcacctc ctgcgatcgc gagccgatcc
     2281 acatccccgg cagcattcag ccgtgcggct gcctgctagc ctgcgacgcg caggcggtgc
     2341 ggatcacgcg cattacggaa aatgccggcg cgttctttgg acgcgaaact ccgcgggtcg
     2401 gtgagctact cgccgattac ttcggcgaga ccgaagccca tgcgctgcgc aacgcactgg
     2461 cgcagtcctc cgatccaaag cgaccggcgc tgatcttcgg ttggcgcgac ggcctgaccg
     2521 gccgcacctt cgacatctca ctgcatcgcc atgacggtac atcgatcatc gagttcgagc
     2581 ctgcggcggc cgaacaggcc gacaatccgc tgcggctgac gcggcagatc atcgcgcgca
     2641 ccaaagaact gaagtcgctc gaagagatgg ccgcacgggt gccgcgctat ctgcaggcga
     2701 tgctcggcta tcaccgcgtg atgttgtacc gcttcgcgga cgacggctcc gggatggtga
     2761 tcggcgaggc gaagcgcagc gacctggaga gctttctcgg tcagcacttt ccggcgtcgc
     2821 tggtcccgca gcaggcgcgg ctactgtact tgaagaacgc gatccgcgtg gtctcggatt
     2881 cgcgcggcat cagcagccgg atcgtgcccg agcacgacgc ctccggcgcc gcgctcgatc
     2941 tgtcgttcgc gcacctgcgc agcatctcgc cctgccatct cgaatttctg cggaacatgg
     3001 gcgtcagcgc ctcgatgtcg ctgtcgatca tcattgacgg cacgctatgg ggattgatca
     3061 tctgtcatca ttacgagccg cgtgccgtgc cgatggcgca gcgcgtcgcg gccgaaatgt
     3121 tcgccgactt cttatcgctg cacttcaccg ccgcccacca ccaacgctga ttaattaaga
     3181 attcgaccca gctttcttgt acaaagtggt tgatatccag cacagtggcg gccgctcgag
     3241 tctagagggc ccgcggttcg aaggtaagcc tatccctaac cctctcctcg gtctcgattc
     3301 tacgcgtacc ggttagcaat tgttttttcg atgagtttgg acaaaccaca actagaatgc
     3361 agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
     3421 taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg
     3481 gggaggtgtg ggaggttttt taaagcaagt aaaacctcta caaatgtggt acttaagagg
     3541 gggagaccaa agggcgagac gttaaggcct cacgtgacat gtgagcaaaa ggccagcaaa
     3601 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     3661 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     3721 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     3781 ttacgggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     3841 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     3901 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     3961 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     4021 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     4081 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     4141 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     4201 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     4261 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gccgtctcag aagaactcgt
     4321 caagaaggcg atagaaggcg atgcgctgcg aatcgggagc ggcgataccg taaagcacga
     4381 ggaagcggtc agcccattcg ccgccaagct cttcagcaat atcacgggta gccaacgcta
     4441 tgtcctgata gcggtccgcc acacccagcc ggccacagtc gatgaatcca gaaaagcggc
     4501 cattttccac catgatattc ggcaagcagg catcgccatg ggtcacgacg agatcctcgc
     4561 cgtcgggcat gctcgccttg agcctggcga acagttcggc tggcgcgagc ccctgatgct
     4621 cttcgtccag atcatcctga tcgacaagac cggcttccat ccgagtacgt gctctctcga
     4681 tgcgatgttt cgcttggtgg tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc
     4741 gcattgcatc agccatgatg gatactttct cggcaggagc aaggtgagat gacaggagat
     4801 cctgccccgg cacttcgccc aatagcagcc agtcccttcc cgcttcagtg acaacgtcga
     4861 gtacagctgc gcaaggaacg cccgtcgtgg ccagccacga tagccgcgct gcctcgtctt
     4921 gcagttcatt cagggcaccg gacaggtcgg tcttgacaaa aagaaccggg cgcccctgcg
     4981 ctgacagccg gaacacggcg gcatcagagc agccgattgt ctgttgtgcc cagtcatagc
     5041 cgaatagcct ctccacccaa gcggccggag aacctgcgtg caatccatct tgttcaatca
     5101 taatattatt gaagcattta tcagggttcg tctcgtcccg gtctcctccc atgcatg
//
