LOCUS       sequence-242326-        3675 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cloning vector for tRNA-released synergistic activation mediator 
            gRNA. BsmBI flanked placeholder for spacer cloning..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3675)
  AUTHORS   Knapp DJHF, Michaels YS, Jamilly M, Ferry QRV, Barbosa H, Milne TA, 
            Fulga TA
  TITLE     Decoupling tRNA promoter and processing activities enables specific 
            Pol-II Cas9 guide RNA expression.
  JOURNAL   Nat Commun. 2019 Apr 2;10(1):1490. doi: 10.1038/s41467-019-09148-3.
  PUBMED    30940799
REFERENCE   2  (bases 1 to 3675)
  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: "Nat 
            Commun."; date: "2019-04-2"; volume: "10(1):1490. doi"; pages: " 
            10.1038/s41467-019-09148-3"
COMMENT     Sequence Label: 8xCTS1_WT-hutRNApro_BsmBIplch-SAMsgRNA_14nt
            Bu...
FEATURES             Location/Qualifiers
     source          1..3675
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    193..314
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(230..249)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     284..303
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_RNA        complement(461..479)
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        complement(531..549)
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_feature    1101..1192
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     complement(1133..1152)
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     polyA_signal    1206..1254
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(1385..1840)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1522..1543)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1734..1753
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1867..1971
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1972..2040,2041..2832)
                     /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(2190..2209)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3003..3591
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3492..3511
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
       61 tctcaagggc atcggtcgac atactcgatc cttatcgatt ttaccacatt tgtagaggtt
      121 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
      181 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
      241 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
      301 atcaatgtat cttatcatgt ctgctcgaag cggccggccg tcccgggctc gtccgggatt
      361 tgaacccggg acctctcgca ccctaagcga gaatcatacc cctagaccaa cgagccgaca
      421 caaactggtt gcaccgactc ggtgccactt ggccctgcag acatgggtga tcctcatgtt
      481 ggccaagttg ataacggact agccttattt taacttgcta ggccctgcag acatgggtga
      541 tcctcatgtt ggcctagctc taaaacggag acgtccgtct ccgggctcgt ccgggatttg
      601 aacccgggac ctctcgcacc ctaagcgaga atcatacccc tagaccaacg agccgttggc
      661 atggtcaacg cgacgctaga tagcaggacg aacgccccca ccccctttta tagccccccg
      721 ctagcctgaa aatattaata gtcgcgtgta gcgaagcatg gctagcctga aaatattaat
      781 agtcgcgtgt agcgaagcat ggctagcctg aaaatattaa tagtcgcgtg tagcgaagca
      841 tggctagcct gaaaatatta atagtcgcgt gtagcgaagc atgactagcc tgaaaatatt
      901 aatagtcgcg ttagcgaagc atggctagcc tgaaaatatt aatagtcgcg tgtagcgaag
      961 catggctagc ttgaaaatat taatagtcgc gtgtagcgaa gcatggctag cctgaaaata
     1021 ttaatagtcg cgtgtagcga agcatggcta gcctgaaaat attaatagtc gcgacattaa
     1081 tatttcagct agctatcgat agagaaatgt tctggcacct gcacttgcac tggggacagc
     1141 ctattttgct agtttgtttt gtttcgtttt gttttgatgg agagcgtatg ttagtactat
     1201 cgattcacac aaaaaaccaa cacacagatg taatgaaaat aaagatattt tattgcggcc
     1261 gctccaagta cctcccgtac cttaatatta cttacttatc atggtagctt gggctggcgt
     1321 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     1381 tggcaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     1441 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     1501 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     1561 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     1621 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     1681 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     1741 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     1801 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga
     1861 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     1921 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     1981 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     2041 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     2101 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     2161 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     2221 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     2281 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     2341 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     2401 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     2461 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     2521 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     2581 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     2641 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     2701 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     2761 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     2821 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     2881 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     2941 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     3001 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     3061 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     3121 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     3181 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     3241 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     3301 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     3361 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     3421 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     3481 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     3541 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     3601 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gcacctgtcc
     3661 tacgagttgc atcat
//
