LOCUS       sequence-319295-        3279 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tRNA and scaffold 2.1scRNA aptamer Ms2 for the assembly of 
            GBoligomers for the first position (positon [M2_3]) of a 
            polycistronic tRNA-gRNA regulated by the U6-26 or U6-1 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3279)
  AUTHORS   Selma S, Bernabe-Orts JM, Vazquez-Vilar M, Diego-Martin B, Ajenjo M,
            Garcia-Carpintero V, Granell A, Orzaez D
  TITLE     Strong gene activation in plants with genome-wide specificity using 
            a new orthogonal CRISPR/Cas9-based programmable transcriptional 
            activator.
  JOURNAL   Plant Biotechnol J. 2019 Sep;17(9):1703-1705. doi: 
            10.1111/pbi.13138. Epub 2019 May 23.
  PUBMED    31034138
REFERENCE   2  (bases 1 to 3279)
  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: "10.1111/pbi.13138";
            journalName: "Plant Biotechnol J"; date: "2019-09"; volume: "17"; 
            issue: "9"; pages: "1703-1705"
COMMENT     Sequence Label: pVD1 M2_3-pTRNA-scf 2.1 (GB2074)
FEATURES             Location/Qualifiers
     source          1..3279
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     411..430
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     664..681
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    798..819
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(834..839,840..857,858..864)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    872..888
                     /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)."
     primer_bind     877..899
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     896..912
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     896..912
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        930..948
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     930..947
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     misc_RNA        complement(1100..1175)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     regulatory      1306..1315
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1310..1319
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1349..1368)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1350..1368)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1375..1392)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1375..1391)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1384..1406)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1532..1987
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1619..1638)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1829..1850
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2065..2169
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2170..2238,2239..3030)
                     /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(2388..2407)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(3201..3279,1..510)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
       61 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
      121 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
      181 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
      241 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
      301 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
      361 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
      421 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
      481 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
      541 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
      601 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
      661 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
      721 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
      781 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
      841 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
      901 aacagctatg accatgatta cgccaagcta tttaggtgac actatagaat actcaagcta
      961 tgcatccaac gcgttgggag ctctcccata tggtcgacct gcaggcggcc gcgaattcac
     1021 tagtgatccc gtctcgctca ttgggaagac tccccagtga ctgtgggagt cgcacatggg
     1081 tgattcctca tgtgctcccg caccgactcg gtgccacttt ttcaagttga taacggacta
     1141 gccttatttt aacttgctat ttctagctct aaaactgaga cggcgcgcgc cgtctctgca
     1201 ccagccggga atcgaacccg ggtctgtacc gtggcagggt actattctac cactagacca
     1261 ctggtgcttt gttgggacga gcgagacggg aatcgaattc ccgcggccgc catggcggcc
     1321 gggagcatgc gacgtcgggc ccaattcgcc ctatagtgag tcgtattaca attcactggc
     1381 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc
     1441 agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc
     1501 ccaacagttg cgcagcctga atggcgaatg gacgcgccct gtagcggcgc attaagcgcg
     1561 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     1621 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     1681 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     1741 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     1801 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     1861 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     1921 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt
     1981 acaatttcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatca
     2041 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     2101 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     2161 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     2221 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     2281 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     2341 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     2401 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     2461 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     2521 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     2581 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     2641 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     2701 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     2761 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     2821 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     2881 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     2941 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     3001 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     3061 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     3121 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     3181 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3241 acaaaaaaac caccgctacc agcggtggtt tgtttgccg
//
