LOCUS       sequence-232857-        3128 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression esgRNA in wheat protoplasts.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3128)
  AUTHORS   Li C, Zong Y, Wang Y, Jin S, Zhang D, Song Q, Zhang R, Gao C
  TITLE     Expanded base editing in rice and wheat using a Cas9-adenosine 
            deaminase fusion.
  JOURNAL   Genome Biol. 2018 May 29;19(1):59. doi: 10.1186/s13059-018-1443-z.
  PUBMED    29807545
REFERENCE   2  (bases 1 to 3128)
  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: "2018-05-29"; volume: "19(1):59. doi"; pages: " 
            10.1186/s13059-018-1443-z"
COMMENT     Sequence Label: pTaU6-esgRNA
FEATURES             Location/Qualifiers
     source          1..3128
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     127..149
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     141..158
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     142..158
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      632..651
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     primer_bind     complement(670..686)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(670..686)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(683..705)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    694..710
                     /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(718..724,725..742,743..748))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    763..784
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(901..918)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1072..1660)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1152..1171)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1831..2622,2623..2691))
                     /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     2454..2473
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2692..2796)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2864..2882
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2920..2942)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3042..3061
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc
       61 tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag
      121 ggttttccca gtcacgacgt tgtaaaacga cggccagtga attcgaccaa gcccgttatt
      181 ctgacagttc tggtgctcaa cacatttata tttatcaagg agcacattgt tactcactgc
      241 taggagggaa tcgaactagg aatattgatc agaggaacta cgagagagct gaagataact
      301 gccctctagc tctcactgat ctgggcgcat agtgagatgc agcccacgtg agttcagcaa
      361 cggtctagcg ctgggctttt aggcccgcat gatcgggctt tgtcgggtgg tcgacgtgtt
      421 cacgattggg gagagcaacg cagcagttcc tcttagttta gtcccacctc gcctgtccag
      481 cagagttctg accggtttat aaactcgctt gctgcatcag acttgtgaga ccctgcaggg
      541 tctctgttta agagctatgc tggaaacagc atagcaagtt taaataaggc tagtccgtta
      601 tcaacttgaa aaagtggcac cgagtcggtg ctttttttgt tttttatgtc taagcttggc
      661 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
      721 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
      781 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
      841 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
      901 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
      961 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     1021 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1081 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1141 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1201 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1261 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1321 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1381 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     1441 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     1501 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     1561 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     1621 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     1681 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     1741 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     1801 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     1861 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     1921 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagaaccacg
     1981 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     2041 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     2101 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     2161 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     2221 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     2281 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     2341 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     2401 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     2461 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     2521 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     2581 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     2641 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     2701 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     2761 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     2821 tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag
     2881 gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     2941 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     3001 gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt
     3061 actgagagtg caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     3121 catcaggc
//
