LOCUS       sequence-242309-        3613 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 3613)
  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 3613)
  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_deltaC55G-hutRNApro_BsmBIplch-SAMsgRNA_...
FEATURES             Location/Qualifiers
     source          1..3613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(429..1220,1221..1289))
                     /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     1052..1071
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1290..1394)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1421..1876
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1508..1527)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1718..1739
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    2007..2055
                     /note="synthetic polyadenylation signal"
     misc_feature    2069..2160
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     2109..2128
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     misc_RNA        2681..2699
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        2751..2769
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     polyA_signal    2885..3006
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2896..2915)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2950..2969
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(join(3283..3613,1..258))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3363..3382)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
       61 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
      121 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
      181 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
      241 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
      301 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
      361 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
      421 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
      481 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
      541 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
      601 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
      661 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
      721 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
      781 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
      841 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
      901 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
      961 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     1021 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     1081 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     1141 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     1201 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     1261 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     1321 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     1381 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgcgccctg tagcggcgca
     1441 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     1501 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     1561 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     1621 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     1681 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     1741 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     1801 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     1861 ttaacgctta caatttgcca ttcgccattc aggctgcgca actgttggga agggcgatcg
     1921 gtgcgggcct cttcgctatt acgccagccc aagctaccat gataagtaag taatattaag
     1981 gtacgggagg tacttggagc ggccgcaata aaatatcttt attttcatta catctgtgtg
     2041 ttggtttttt gtgtgaatcg atagtactaa catacgctct ccatcaaaac aaaacgaaac
     2101 aaaacaaact agcaaaatag gctgtcccca gtgcaagtgc aggtgccaga acatttctct
     2161 atcgatagct agctgaaata ttaatgtcgc gactattaat attttcaggc tagccatgct
     2221 tcgctacacg cgactattaa tattttcagg ctagccatgc ttcgctacac gcgactatta
     2281 atattttcaa gctagccatg cttcgctaca cgcgactatt aatattttca ggctagccat
     2341 gcttcgctaa cgcgactatt aatattttca ggctagtcat gcttcgctac acgcgactat
     2401 taatattttc aggctagcca tgcttcgcta cacgcgacta ttaatatttt caggctagcc
     2461 atgcttcgct acacgcgact attaatattt tcaggctagc catgcttcgc tacacgcgac
     2521 tattaatatt ttcaggctag cggggggcta taaaaggggg tgggggcgtt cgtcctgcta
     2581 tctagcgtcg cgttgaccat gccaacggct cgttgggagg tcccgggttg aaatcccgga
     2641 cgagcccgga gacggacgtc tccgttttag agctaggcca acatgaggat cacccatgtc
     2701 tgcagggcct agcaagttaa aataaggcta gtccgttatc aacttggcca acatgaggat
     2761 cacccatgtc tgcagggcca agtggcaccg agtcggtgca accagtttgt gtcggctcgt
     2821 tgggaggtcc cgggttgaaa tcccggacga gcccgggacg gccggccgct tcgagcagac
     2881 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     2941 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     3001 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag
     3061 gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tcgagtatgt
     3121 cgaccgatgc ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg
     3181 actatcgtcg ccgcacttat gatgcaactc gtaggacagg tgcatgtgag caaaaggcca
     3241 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     3301 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     3361 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     3421 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     3481 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     3541 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3601 cccggtaaga cac
//
