LOCUS       sequence-403361-        6406 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of CRISPR RNA, shuttle vector for Corynebacterium 
            glutamicum / E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6406)
  AUTHORS   Yang S, Zhu J, Zhou X, Zhang J, Li Q, Bian F, Zhu J, Yan T, Wang X, 
            Zhang Y, Yang J, Jiang Y, Yang S
  TITLE     RNA-Guided DNA Transposition in Corynebacterium glutamicum and 
            Bacillus subtilis.
  JOURNAL   ACS Synth Biol. 2023 Jul 21;12(7):2198-2202. doi: 
            10.1021/acssynbio.3c00193. Epub 2023 Jun 22.
  PUBMED    37348112
REFERENCE   2  (bases 1 to 6406)
  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: 
            "10.1021/acssynbio.3c00193"; journalName: "ACS Synth Biol"; date: 
            "2023-07-21- 21"; volume: "12"; issue: "7"; pages: "2198-2202"
COMMENT     Sequence Label: pCgVchcrRNA
FEATURES             Location/Qualifiers
     source          1..6406
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(100..894)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(211..230)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     821..840
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      complement(3853..4591)
                     /direction=LEFT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     terminator      complement(4819..4846)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(4938..5023)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(5104..5727)
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCKSGS"
     primer_bind     5635..5654
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    6147..6165
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
ORIGIN
        1 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
       61 caaatatgta tccgctcatg aattaattca agcttcatat cagaagaact cgtcaagaag
      121 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg
      181 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg
      241 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc
      301 caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct cgccgtcggg
      361 catccgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc
      421 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg
      481 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc
      541 atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc
      601 cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc
      661 tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cttggagttc
      721 attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag
      781 ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag
      841 cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatgcgaaa
      901 cgatcctcat cctgtctctt gatcagatct tgatcccctg cgccatcaga tccttggcgg
      961 caagaaagcc atccagttta ctttgcaggg cttcccaacc ttaccagagg gcgccccagc
     1021 tggcaattcc ggttcgcttg ctgtccataa aaccgcccag tctagctatc gccatgtaag
     1081 cccactgcaa gctacctgct ttctctttgc gcttgcgttt tcccttgtcc agatagccca
     1141 gtagctgaca ttcatccggg gtcagcaccg tttctgcgga ctggctttct acgtgttccg
     1201 cttccggtga ccgcctcgat gatcgccggg tgggcgtggc ccaaggagga tcatccagcc
     1261 attcggggtc gttcactggt tcccctttct gatttctggc atagaagaac ccccgtgaac
     1321 tgtgtggttc cgggggttgc tgatttttgc gagacttctc gcgcaattcc ctagcttagg
     1381 tgaaaacacc atgaaacact agggaaacac ccatgaaaca cccattaggg cagtagggcg
     1441 gcttcttcgt ctagggcttg catttgggcg gtgatctggt ctttagcgtg tgaaagtgtg
     1501 tcgtaggtgg cgtgctcaat gcactcgaac gtcacgtcat ttaccgggtc acggtgggca
     1561 aagagaacta gtgggttaga cattgttttc ctcgttgtcg gtggtggtga gcttttctag
     1621 ccgctcggta aacgcggcga tcatgaactc ttggaggttt tcaccgttct gcatgcctgc
     1681 gcgcttcatg tcctcacgta gtgccaaagg aacgcgtgcg gtgaccacga cgggcttagc
     1741 ctttgcctgc gcttctagtg cttcgatggt ggcttgtgcc tgcgcttgct gcgcctgtag
     1801 tgcctgttga gcttcttgta gttgctgttc tagctgtgcc ttggttgcca tgctttaaga
     1861 ctctagtagc tttcctgcga tatgtcatgc gcatgcgtag caaacattgt cctgcaactc
     1921 attcattatg tgcagtgctc ctgttactag tcgtacatac tcatatttac ctagtctgca
     1981 tgcagtgcat gcacatgcag tcatgtcgtg ctaatgtgta aaacatgtac atgcagattg
     2041 ctgggggtgc agggggcgga gccaccctgt ccatgcgggg tgtggggctt gccccgccgg
     2101 tacagacagt gagcaccggg gcacctagtc gcggataccc cccctaggta tcggacacgt
     2161 aaccctccca tgtcgatgca aatctttaac attgagtacg ggtaagctgg cacgcatagc
     2221 caagctaggc ggccaccaaa caccactaaa aattaatagt ccctagacaa gacaaacccc
     2281 cgtgcgagct accaactcat atgcacgggg gccacataac ccgaaggggt ttcaattgac
     2341 aaccatagca ctagctaaga caacgggcac aacacccgca caaactcgca ctgcgcaacc
     2401 ccgcacaaca tcgggtctag gtaacactga aatagaagtg aacacctcta aggaaccgca
     2461 ggtcaatgag ggttctaagg tcactcgcgc tagggcgtgg cgtaggcaaa acgtcatgta
     2521 caagatcacc aatagtaagg ctctggcggg gtgccatagg tggcgcaggg acgaagctgt
     2581 tgcggtgtcc tggtcgtcta acggtgcttc gcagtttgag ggtctgcaaa actctcactc
     2641 tcgctggggg tcatctctgg ctgaattgga agtcatgggc gaacgccgca ttgagctggc
     2701 tattgctact aagaatcact tggcggcggg tggcgcgctc atgatgtttg tgggcactgt
     2761 tcgacacaac cgctcacagt catttgcgca ggttgaagcg ggtattaaga ctgcgtactc
     2821 ttcgatggtg aaaacatctc agtggaagaa agaacgtgca cggtacgggg tggagcacac
     2881 ctatagtgac tatgaggtca cagactcttg ggcgaacggt tggcacttgc accgcaacat
     2941 gctgttgttc ttggatcgtc cactgtctga cgatgaactc aaggcgtttg aggattccat
     3001 gttttcccgc tggtctgctg gtgtggttaa ggccggtatg gacgcgccac tgcgtgagca
     3061 cggggtcaaa cttgatcagg tgtctacctg gggtggagac gctgcgaaaa tggcaaccta
     3121 cctcgctaag ggcatgtctc aggaactgac tggctccgct actaaaaccg cgtctaaggg
     3181 gtcgtacacg ccgtttcaga tgttggatat gttggccgat caaagcgacg ccggcgagga
     3241 tatggacgct gttttggtgg ctcggtggcg tgagtatgag gttggttcta aaaacctgcg
     3301 ttcgtcctgg tcacgtgggg ctaagcgtgc tttgggcatt gattacatag acgctgatgt
     3361 acgtcgtgaa atggaagaag aactgtacaa gctcgccggt ctggaagcac cggaacgggt
     3421 cgaatcaacc cgcgttgctg ttgctttggt gaagcccgat gattggaaac tgattcagtc
     3481 tgatttcgcg gttaggcagt acgttctcga ttgcgtggat aaggctaagg acgtggccgc
     3541 tgcgcaacgt gtcgctaatg aggtgctggc aagtctgggt gtggattcca ccccgtgcat
     3601 gatcgttatg gatgatgtgg acttggacgc ggttctgcct actcatgggg acgctactaa
     3661 gcgtgatctg aatgcggcgg tgttcgcggg taatgagcag actattcttc gcacccacta
     3721 aaagcggcat aaaccccgtt cgatattttg tgcgatgaat ttatggtcaa tgtcgcgggg
     3781 gcaaactatg atgggtcttg ttgtttagct agctcactcg gtcgctacgc tccgggcgtg
     3841 agactgcggc gggcgctgcg gacacataca aagttaccca cagattccgt ggataagcag
     3901 gggactaaca tgtgaggcaa aacagcaggg ccgcgccggt ggcgtttttc cataggctcc
     3961 gccctcctgc cagagttcac ataaacagac gcttttccgg tgcatctgtg ggagccgtga
     4021 ggctcaacca tgaatctgac agtacgggcg aaacccgaca ggacttaaag atccccaccg
     4081 tttccggcgg gtcgctccct cttgcgctct cctgttccga ccctgccgtt taccggatac
     4141 ctgttccgcc tttctccctt acgggaagtg tggcgctttc tcatagctca cacactggta
     4201 tctcggctcg gtgtaggtcg ttcgctccaa gctgggctgt aagcaagaac tccccgttca
     4261 gcccgactgc tgcgccttat ccggtaactg ttcacttgag tccaacccgg aaaagcacgg
     4321 taaaacgcca ctggcagcag ccattggtaa ctgggagttc gcagaggatt tgtttagcta
     4381 aacacgcggt tgctcttgaa gtgtgcgcca aagtccggct acactggaag gacagatttg
     4441 gttgctgtgc tctgcgaaag ccagttacca cggttaagca gttccccaac tgacttaacc
     4501 ttcgatcaaa ccacctcccc aggtggtttt ttcgtttaca gggcaaaaga ttacgcgcag
     4561 aaaaaaagga tctcaagaag atcctttgat cttttctact gaaccgctct agatttcagt
     4621 gcaatttatc tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt
     4681 ctcatgttag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag
     4741 ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc
     4801 agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg ccttctgctt aatttgatgc
     4861 ctggcagttt atggcgggcg tcctgcccgc caccctccgg gccgttgctt cgcaacgttc
     4921 aaatccgctc ccggcggatt tgtcctactc aggagagcgt tcaccgacaa acaacagata
     4981 aaacgaaagg cccagtcttt cgactgagcc tttcgtttta tttaaattct agactgatga
     5041 atcccctaat gatttttatc aaaatcatta aggttaccat cacggaaaaa ggttatgctg
     5101 cttttaagac ccacttttac atttaagttg tttttctaat ccgcatatga tcaattcaag
     5161 gccgaataag aaggctggct ctgcaccttg gtgatcaaat aattcgatag cttgtcgtaa
     5221 taatggcggc atactatcag tagtaggtgt ttccctttct tctttagcga cttgatgctc
     5281 ttgatcttcc aatacgcaac ctaaagtaaa atgccccaca gcgctgagtg catataatgc
     5341 attctctagt gaaaaacctt gttggcataa aaaggctaat tgattttcga gagtttcata
     5401 ctgtttttct gtaggccgtg tacctaaatg tacttttgct ccatcgcgat gacttagtaa
     5461 agcacatcta aaacttttag cgttattacg taaaaaatct tgccagcttt ccccttctaa
     5521 agggcaaaag tgagtatggt gcctatctaa catctcaatg gctaaggcgt cgagcaaagc
     5581 ccgcttattt tttacatgcc aatacaatgt aggctgctct acacctagct tctgggcgag
     5641 tttacgggtt gttaaacctt cgattccgac ctcattaagc agctctaatg cgctgttaat
     5701 cactttactt ttatctaatc tagacatgtt gtgtctcctc taaagattgt aggaaatgca
     5761 atgtgtcaag cagttaggtc aatcagagca tcggtcactc aagaattctt gagctgacag
     5821 aacccaaggg cgagtatggc gaatttttcc aactcacgca ccaaacctac tcacattgga
     5881 taattcgtcg agtcaactcc gtgtcaacct gtcctcaact ttagcgggga aacaaagatt
     5941 tcgcagcaga tgcaaaatca tacagacaca ttcaaaaatg gacattgacc ccaaacgggg
     6001 gcatttccaa cacttgcgct aaatagtttt tgcagggaac gaacctgaaa tcgttcgcac
     6061 tacgatctgc tcccccgaat ggggttaatt atgaactaca ttacaccccc gtgaatcgaa
     6121 ttccaaaaac taaaaaaaat attgacactc tatcattgat agagtataat taacgggatc
     6181 cactcttcat tgataagtcg ccaagcttgg gatccccagc ttgttgatac actaatgctt
     6241 ttatataggg aaaaggtggt gaactactcc atgggtgaac tgccgagtag gtagctgata
     6301 acgagacctc tggtctcgtg aactgccgag taggtagctg ataacggatc cgaattcgag
     6361 ctcggcgcgc ctgcaggtcg acaagcttgc gctcgagcgg gggccc
//
