LOCUS       sequence-257378-        3182 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses gRNA for SLBP-based CIRTS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3182)
  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 3182)
  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: SLBP non-targeting gRNA lambda phage
FEATURES             Location/Qualifiers
     source          1..3182
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     47..69
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     61..78
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     62..78
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     88..107
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        88..106
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        121..369
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     121..141
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(456..476)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(456..474)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(495..511)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(495..511)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(508..530)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    519..535
                     /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(543..549,550..567,568..573))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    588..609
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(726..743)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(897..1485)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(977..996)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1656..2447,2448..2516))
                     /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     2279..2298
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2517..2621)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2647..3102)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2784..2805)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
       61 tgtaaaacga cggccagtga gcgcgcgtaa tacgactcac tatagggcga attgggtacc
      121 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
      181 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      241 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      301 atgcttatcg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      361 cgaaacaccg gccaaaggct cttctcagag ccacccatta ttgtgataag tggaatgcca
      421 tgtttttttc cgcggtggag ctccagcttt tgttcccttt agtgagggtt aattgcgcgc
      481 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
      541 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
      601 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
      661 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
      721 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
      781 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
      841 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
      901 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
      961 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     1021 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     1081 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     1141 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     1201 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     1261 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     1321 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     1381 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     1441 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     1501 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     1561 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     1621 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     1681 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     1741 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     1801 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     1861 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     1921 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     1981 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     2041 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     2101 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     2161 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     2221 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     2281 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     2341 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     2401 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     2461 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     2521 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     2581 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     2641 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     2701 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     2761 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     2821 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     2881 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     2941 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaagtga
     3001 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     3061 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc
     3121 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     3181 aa
//
