LOCUS       sequence-408849-        3482 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For plant prime editing in wheat plants or monocotyledons 
            protoplasts.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3482)
  AUTHORS   Ni P, Zhao Y, Zhou X, Liu Z, Huang Z, Ni Z, Sun Q, Zong Y
  TITLE     Efficient and versatile multiplex prime editing in hexaploid wheat.
  JOURNAL   Genome Biol. 2023 Jun 29;24(1):156. doi: 10.1186/s13059-023-02990-1.
  PUBMED    37386475
REFERENCE   2  (bases 1 to 3482)
  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: "Genome 
            Biol."; date: "2023-06-29"; volume: "24(1):156. doi"; pages: " 
            10.1186/s13059-023-02990-1"
COMMENT     Sequence Label: pUC57-CmYLCV
FEATURES             Location/Qualifiers
     source          1..3482
                     /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"
     protein_bind    867..886
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     protein_bind    1042..1061
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     polyA_signal    1068..1242
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     primer_bind     complement(1261..1277)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1261..1277)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1274..1296)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1285..1301
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1309..1315,1316..1333,1334..1339))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1354..1375
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1492..1509)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1663..2251)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1743..1762)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2422..3213,3214..3282))
                     /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     3045..3064
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3283..3387)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3455..3473
                     /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 ctggcagaca tactgtccca
      421 caaatgaaga tggaatctgt aaaagaaaac gcgtgaaata atgcgtctga caaaggttag
      481 gtcggctgcc tttaatcaat accaaagtgg tccctaccac gatggaaaaa ctgtgcagtc
      541 ggtttggctt tttctgacga acaaataaga ttcgtggccg acaggtgggg gtccaccatg
      601 tgaaggcatc ttcagactcc aataatggag caatgacgta agggcttacg aaataagtaa
      661 gggtagtttg ggaaatgtcc actcacccgt cagtctataa atacttagcc cctccctcat
      721 tgttaaggga gcaaaatctc agagagatag tcctagagag agaaagagag caagtagcct
      781 agaagtagtc aaggcggcga agtattcagg cacgtggcca ggaagaagaa aagccaagac
      841 gacgaaaaca ggtaagagct aagcttgttc actgccgtat aggcagcgag accaaggtct
      901 cagtttaaga gctatgctgg aaacagcata gcaagtttaa ataaggctag tccgttatca
      961 acttgaaaaa gtggcaccga gtcggtgcgg agaccaaggt ctcccgcggt tctatctagt
     1021 tacgcgttaa accaactaga agttcactgc cgtataggca gggatccttt ctccataata
     1081 atgtgtgagt agttcccaga taagggaatt agggttccta tagggtttcg ctcatgtgtt
     1141 gagcatataa gaaaccctta gtatgtattt gtatttgtaa aatacttcta tcaataaaat
     1201 ttctaattcc taaaaccaaa atccagtact aaaatccaga tcaagcttgg cgtaatcatg
     1261 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     1321 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     1381 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     1441 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     1501 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     1561 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     1621 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     1681 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     1741 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     1801 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     1861 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     1921 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     1981 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     2041 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     2101 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     2161 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     2221 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     2281 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     2341 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     2401 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     2461 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     2521 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagaaccac gctcaccggc
     2581 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     2641 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     2701 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     2761 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     2821 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     2881 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     2941 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     3001 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     3061 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     3121 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     3181 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     3241 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     3301 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     3361 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     3421 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     3481 tc
//
