LOCUS       sequence-254557-        3206 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses gRNA for TBP-based CIRTS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3206)
  AUTHORS   Rauch S, He E, Srienc M, Zhou H, Zhang Z, Dickinson BC
  TITLE     Programmable RNA-Guided RNA Effector Proteins Built from Human 
            Parts.
  JOURNAL   Cell. 2019 Jun 27;178(1):122-134.e12. doi: 
            10.1016/j.cell.2019.05.049. Epub 2019 Jun 20.
  PUBMED    31230714
REFERENCE   2  (bases 1 to 3206)
  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.2019.05.049"; journalName: "Cell"; date: 
            "2019-06-27- 27"; volume: "178"; issue: "1"; pages: "122-134.e12"
COMMENT     Sequence Label: TBP6.7 non-targeting gRNA lambda phage
FEATURES             Location/Qualifiers
     source          1..3206
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        659..907
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     659..679
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(1018..1038)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1018..1036)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1057..1073)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1057..1073)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1070..1092)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1081..1097
                     /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(1105..1111,1112..1129,1130..1135))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1150..1171
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1288..1305)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1459..2047)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1539..1558)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2218..3009,3010..3078))
                     /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     2841..2860
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3079..3183)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtaccga
      661 gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg ttagagagat
      721 aattggaatt aatttgactg taaacacaaa gatattagta caaaatacgt gacgtagaaa
      781 gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg actatcatat
      841 gcttatcgta acttgaaagt atttcgattt cttggcttta tatatcttgt ggaaaggacg
      901 aaacaccggg ccagatctga gcctgggagc tctctggcct tatttgacag cccacatggc
      961 attccactta tcactggcat cctttttttt tccgcggtgg agctccagct tttgttccct
     1021 ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     1081 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     1141 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     1201 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     1261 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     1321 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     1381 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     1441 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     1501 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     1561 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     1621 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     1681 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     1741 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     1801 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     1861 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     1921 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     1981 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     2041 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     2101 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     2161 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     2221 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     2281 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     2341 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     2401 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     2461 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     2521 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     2581 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     2641 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     2701 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     2761 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     2821 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     2881 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     2941 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     3001 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     3061 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     3121 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     3181 gcgcacattt ccccgaaaag tgccac
//
