LOCUS       sequence-292704-        2195 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Entry cloning vector for in vitro transcription or expression of 
            SpCas9 sgRNAs from a T7 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2195)
  TITLE     Joung Lab Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 2195)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pT7-[BsaI_cassette]-SpCas9_sgRNA_scaffold (MSP...
FEATURES             Location/Qualifiers
     source          1..2195
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_RNA        89..164
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(346..363)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(346..362)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      403..990
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     891..910
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1002..1029)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(1053..1868)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1776..1795
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(1869..1971)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(1972..2003)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     primer_bind     2051..2067
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2051..2067
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 taatacgact cactataggg agacccatgc catagcgttg ttcggaatat gaatttttga
       61 acagattcac caacacctag tggtctcagt tttagagcta gaaatagcaa gttaaaataa
      121 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgcttttaa aaagcttgga
      181 tcgacgagag cagcgcgact ggatctgtcg cccgtctcaa acgcaaccct ccggcggtcg
      241 catatcattc aggacgagcc tcagactcca gcgtaactgg actgcaatca actcactggc
      301 tcaccttccg gtccacgatc agctagaatc aagctgacta gataaactgg ccgtcgtttt
      361 acacgggtgg gcctttcttc ggtagaaaat caaaggatct tcttgagatc ctttttttct
      421 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
      481 ggatcaagag ctaccaactc tttttccgag gtaactggct tcagcagagc gcagatacca
      541 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
      601 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
      661 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
      721 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
      781 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
      841 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
      901 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcatcg atttttgtga
      961 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcagaaag gcccacccga
     1021 aggtgagcca ggtgattaca tttaggtcct cattagaaaa actcatcgag catcaagtga
     1081 aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt
     1141 aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct
     1201 gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg
     1261 ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg caagagttta
     1321 tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc
     1381 gcaccaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg
     1441 ccgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc
     1501 gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc
     1561 cctgggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg
     1621 gtcggaagag gcataaattc cgtcagccag tttagcctga ccatctcatc tgtaacatca
     1681 ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac
     1741 aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat
     1801 aaatcagcat ccatgttgga atttaatcgc ggcttcaagc aagacgtttc ccgttgaata
     1861 tggctcattt tagcttcctt agctcctgaa aatctcgata actcaaaaaa tacgcccggt
     1921 agtgatctta tttcattatg gtgaaagttg gaacctctta cgtgccgatc aagtcaaaag
     1981 cctccggtcg gaggcttttg actttctgct atggaggtca ggtatgattt aaatggtcag
     2041 tattgagcct caggaaacag ctatgacatc aagctgacta gataatctag ctgatcgtgg
     2101 accgatcata cgtataatgc cgtaagatca cgggtcgcag cacagctcgc ggtccagtag
     2161 tgatcgacac tgctcgatcc gctcgcaccg ctagc
//
