LOCUS       sequence-339404-        2989 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To construct expression vector for tBE-V5-mA3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2989)
  AUTHORS   Wang L, Xue W, Zhang H, Gao R, Qiu H, Wei J, Zhou L, Lei YN, Wu X, 
            Li X, Liu C, Wu J, Chen Q, Ma H, Huang X, Cai C, Zhang Y, Yang B, 
            Yin H, Yang L, Chen J
  TITLE     Eliminating base-editor-induced genome-wide and transcriptome-wide 
            off-target mutations.
  JOURNAL   Nat Cell Biol. 2021 May 10. pii: 10.1038/s41556-021-00671-4. doi: 
            10.1038/s41556-021-00671-4.
  PUBMED    33972728
REFERENCE   2  (bases 1 to 2989)
  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 Cell 
            Biol. 2021 May 10. pii: 10.1038/s41556-021-00671-4. doi: 
            10.1038/s41556-021-00671-4."
COMMENT     Sequence Label: pUC57-sgRNA-MS2-U6
FEATURES             Location/Qualifiers
     source          1..2989
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        442..460
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_RNA        499..517
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     promoter        628..876
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     628..648
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     799..818
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     protein_bind    905..926
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1044..1061)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1215..1803)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1295..1314)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1974..2789)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1974..2789)
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2697..2716
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2790..2894)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2962..2980
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acctcgcgaa
      421 tgcatctaga tatcggatcc ctaatacgac tcactatagg tgagaccgag agagggtctc
      481 agtttgagag ctaggccaac atgaggatca cccatgtctg cagggcctag caagttcaaa
      541 taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt ttttaaaggg
      601 cccgtcgact gcagaggcta gctcaccgag ggcctatttc ccatgattcc ttcatatttg
      661 catatacgat acaaggctgt tagagagata attggaatta atttgactgt aaacacaaag
      721 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta
      781 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc
      841 ttggctttat atatcttgtg gaaaggacga aacaccgcat aaagtgtaaa gcctggggtg
      901 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
      961 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     1021 gtattgggcg cggccgccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     1081 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     1141 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     1201 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     1261 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     1321 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     1381 ctcccttctg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     1441 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     1501 gccttatccg gtaactatcg tcttgagtcc aacccggcaa gacacgactt atcgccactg
     1561 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     1621 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     1681 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     1741 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     1801 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     1861 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     1921 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttagaaa
     1981 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     2041 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     2101 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     2161 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     2221 ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta
     2281 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     2341 gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     2401 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     2461 aatacctgga atgctgtttt cccagggatc gcagtggtga gtaaccatgc atcatcagga
     2521 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     2581 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     2641 ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg
     2701 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag
     2761 caagacgttt cccgttgaat atggctcata ctcttccttt ttcaatatta ttgaagcatt
     2821 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     2881 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     2941 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtc
//
