LOCUS       sequence-434027-        3685 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Guide RNA expression vector with the synthetic crRNA crR2 targeting 
            the SWITCHER construct with the target site TR2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3685)
  AUTHORS   Schuster B, Dobiasovska I, Curcic J, Pajer P, Borna S, Bartunek P
  TITLE     SWITCHER, a CRISPR-inducible floxed wild-type Cre regulating CRISPR 
            activity.
  JOURNAL   Commun Biol. 2025 Jul 2;8(1):982. doi: 10.1038/s42003-025-08427-8.
  PUBMED    40596718
REFERENCE   2  (bases 1 to 3685)
  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: "Commun 
            Biol."; date: "2025-07-2"; volume: "8(1):982. doi"; pages: " 
            10.1038/s42003-025-08427-8"
COMMENT     Sequence Label: pU6-LbCpf1-crR2
FEATURES             Location/Qualifiers
     source          1..3685
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        301..376
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     polyA_signal    537..585
                     /note="synthetic polyadenylation signal"
     misc_feature    599..690
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     639..658
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     primer_bind     complement(726..743)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    732..939
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   948..1088
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   948..1077
                     /label=AAV2 ITR
     rep_origin      1163..1618
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1250..1269)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1460..1481
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1635..1654)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1754..1776
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1814..1832)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1900..2004
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2005..2073,2074..2865)
                     /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     complement(2223..2242)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3036..3624
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3525..3544
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg taatttctac taagtgtaga tccaacagta ccggattgcc aagctttttt
      301 gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt
      361 ggcaccgagt cggtgctttt ttgttttaga gctagaaata gcaagttaaa ataaggctag
      421 tccgttttta gcgcgtgcgc caattctgca gacaaatggc tctagaggta cctactagta
      481 cgtctctcaa ggataagtaa gtaatattaa ggtacgggag gtattggaca ggccgcaata
      541 aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagtactaa
      601 catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag gctgtcccca
      661 gtgcaagtgc aggtgccaga acatttctct ggcctaactg gccggtacct gagctcgctg
      721 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
      781 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
      841 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
      901 gggggaggat tgggaagaga atagcaggca tgctggggag cggccgcagg aacccctagt
      961 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     1021 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     1081 cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     1141 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     1201 gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct
     1261 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     1321 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     1381 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     1441 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc
     1501 tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat
     1561 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     1621 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     1681 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     1741 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     1801 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     1861 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     1921 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     1981 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     2041 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     2101 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     2161 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     2221 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     2281 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     2341 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     2401 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     2461 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     2521 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     2581 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     2641 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     2701 tctggagccg gtgagcgtgg aagccgcggt atcattgcag cactggggcc agatggtaag
     2761 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     2821 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     2881 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     2941 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     3001 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     3061 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     3121 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     3181 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     3241 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     3301 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     3361 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     3421 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     3481 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     3541 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     3601 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     3661 ttttgctggc cttttgctca catgt
//
