LOCUS       sequence-190277-        3738 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgRNA-FANCF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3738)
  AUTHORS   Tang W, Hu JH, Liu DR
  TITLE     Aptazyme-embedded guide RNAs enable ligand-responsive genome editing
            and transcriptional activation.
  JOURNAL   Nat Commun. 2017 Jun 28;8:15939. doi: 10.1038/ncomms15939.
  PUBMED    28656978
REFERENCE   2  (bases 1 to 3738)
  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: "2017-06-28"; volume: "8:15939. doi"; pages: " 
            10.1038/ncomms15939"
COMMENT     Sequence Label: pWT055f
FEATURES             Location/Qualifiers
     source          1..3738
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        82..330
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     82..102
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     253..272
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        351..426
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        490..589
                     /label=CMVd1 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate 
                     early promoter that yields slightly reduced expression 
                     relative to the full-length promoter"
     primer_bind     540..560
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    612..692
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      694..703
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             700..1635
                     /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"
     polyA_signal    1757..1878
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1794..1813)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1848..1867
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1989..2577)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2069..2088)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2748..3539,3540..3608))
                     /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     3371..3390
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3609..3713)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgtcgcta gctgtacaaa aaagcaggct ttaaaggaac caattcagtc gactggatcc
       61 ggtaccaagg tcgggcagga agagggccta tttcccatga ttccttcata tttgcatata
      121 cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac aaagatatta
      181 gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta
      241 tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct
      301 ttatatatct tgtggaaagg acgaaacacc ggaatccctt ctgcagcacc gttttagagc
      361 tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt
      421 cggtgctttt tttaagcttg ggccgctcga ggtacctctc tacatatgac atgggagttt
      481 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
      541 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa
      601 ccgtcagatc cgctagcgct accggactca gatctcgagc tcaagcttcg aattctgcag
      661 tcgacggtac cgcgggcccg ggatccaccg gtcgccacca tggcgcgtaa ggtcgatctc
      721 acctcctgcg atcgcgagcc gatccacatc cccggcagca ttcagccgtg cggctgcctg
      781 ctagcctgcg acgcgcaggc ggtgcggatc acgcgcatta cggaaaatgc cggcgcgttc
      841 tttggacgcg aaactccgcg ggtcggtgag ctactcgccg attacttcgg cgagaccgaa
      901 gcccatgcgc tgcgcaacgc actggcgcag tcctccgatc caaagcgacc ggcgctgatc
      961 ttcggttggc gcgacggcct gaccggccgc accttcgaca tctcactgca tcgccatgac
     1021 ggtacatcga tcatcgagtt cgagcctgcg gcggccgaac aggccgacaa tccgctgcgg
     1081 ctgacgcggc agatcatcgc gcgcaccaaa gaactgaagt cgctcgaaga gatggccgca
     1141 cgggtgccgc gctatctgca ggcgatgctc ggctatcacc gcgtgatgtt gtaccgcttc
     1201 gcggacgacg gctccgggat ggtgatcggc gaggcgaagc gcagcgacct cgagagcttt
     1261 ctcggtcagc actttccggc gtcgctggtc ccgcagcagg cgcggctact gtacttgaag
     1321 aacgcgatcc gcgtggtctc ggattcgcgc ggcatcagca gccggatcgt gcccgagcac
     1381 gacgcctccg gcgccgcgct cgatctgtcg ttcgcgcacc tgcgcagcat ctcgccctgc
     1441 catctcgaat ttctgcggaa catgggcgtc agcgcctcga tgtcgctgtc gatcatcatt
     1501 gacggcacgc tatggggatt gatcatctgt catcattacg agccgcgtgc cgtgccgatg
     1561 gcgcagcgcg tcgcggccga aatgttcgcc gacttcttat cgctgcactt caccgccgcc
     1621 caccaccaac gctaagcggc cgcgactcta gatcataatc agccatacca catttgtaga
     1681 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     1741 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     1801 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     1861 actcatcaat gtatcttaag gcgtaaattg taagcgttaa tattttgtta aaattcgcgt
     1921 taaattttta tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     1981 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2041 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2101 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2161 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2221 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2281 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2341 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     2401 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     2461 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     2521 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     2581 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     2641 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     2701 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     2761 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     2821 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     2881 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     2941 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     3001 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     3061 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     3121 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     3181 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     3241 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     3301 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     3361 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     3421 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     3481 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     3541 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     3601 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     3661 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     3721 ccccgaaaag tgccacct
//
