LOCUS       sequence-272095-        2805 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Contains the T7 promoter and gRNA scaffold and is used for the in 
            vitro transcription of gRNAs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2805)
  TITLE     Xie Lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 2805)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pUC19-gRNA
FEATURES             Location/Qualifiers
     source          1..2805
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(34..56)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     156..175
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     369..391
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     383..400
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     384..400
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     423..442
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        423..441
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_RNA        458..533
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(589..605)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(589..605)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(602..624)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    613..629
                     /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(637..643,644..661,662..667))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    682..703
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(820..837)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(991..1579)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1071..1090)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1750..2541,2542..2610))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGTRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2373..2392
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2611..2715)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2783..2801
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
       61 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
      121 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
      181 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
      241 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
      301 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
      361 agggttttcc cagtcacgac gttgtaaaac gacggccagt gaattcgagc tcggtaccaa
      421 gctaatacga ctcactatag gggagaccga ggtctcggtt ttagagctag aaatagcaag
      481 ttaaaataag gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttaaa
      541 ggatcctcta gagtcgacct gcaggcatgc aagcttggcg taatcatggt catagctgtt
      601 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
      661 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
      721 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
      781 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
      841 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
      901 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
      961 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     1021 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     1081 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     1141 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     1201 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     1261 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     1321 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     1381 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     1441 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     1501 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     1561 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     1621 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     1681 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     1741 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     1801 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     1861 atctggcccc agtgctgcaa tgataccgcg agtgccacgc tcaccggctc cagatttatc
     1921 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     1981 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     2041 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     2101 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     2161 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     2221 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     2281 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     2341 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     2401 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     2461 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     2521 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     2581 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     2641 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     2701 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat
     2761 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctt
//
