LOCUS       sequence-431496-        4792 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fragmid fragment: (C' terminus) for prime editing.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4792)
  AUTHORS   McGee AV, Liu YV, Griffith AL, Szegletes ZM, Wen B, Kraus C, Miller 
            NW, Steger RJ, Escude Velasco B, Bosch JA, Zirin JD, Viswanatha R, 
            Sontheimer EJ, Goodale A, Greene MA, Green TM, Doench JG
  TITLE     Modular vector assembly enables rapid assessment of emerging CRISPR 
            technologies.
  JOURNAL   Cell Genom. 2024 Mar 13;4(3):100519. doi: 
            10.1016/j.xgen.2024.100519.
  PUBMED    38484704
REFERENCE   2  (bases 1 to 4792)
  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: "Cell 
            Genom."; date: "2024-03-13"; volume: "4(3):100519. doi"; pages: " 
            10.1016/j.xgen.2024.100519"
COMMENT     Sequence Label: AA058
FEATURES             Location/Qualifiers
     source          1..4792
                     /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"
     CDS             2610..2630
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(2702..2718)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2702..2718)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2715..2737)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2726..2742
                     /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(2750..2756,2757..2774,2775..2780))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2795..2816
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2933..2950)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3104..3692)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3184..3203)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3870..4679)
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4593..4612
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4671..4690
                     /label=pENTR-R
                     /note="pENTR vectors, reverse 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 ccagagaatt cgagctcggt acctcgcgaa
      421 tacatctaga tgaagacggg tccggatcta gcggaggatc ctctggcagc gagacaccag
      481 gaacaagcga gtcagcaaca ccagagagca gtggcggcag cagcggcggc agcagcaccc
      541 taaatataga agatgagtat cggctacatg agacctcaaa agagccagat gtttctctag
      601 ggtccacatg gctgtctgat tttcctcagg cctgggcgga aaccgggggc atgggactgg
      661 cagttcgcca agctcctctg atcatacctc tgaaagcaac ctctaccccc gtgtccataa
      721 aacaataccc catgtcacaa gaagccagac tggggatcaa gccccacata cagagactgt
      781 tggaccaggg aatactggta ccctgccagt ccccctggaa cacgcccctg ctacccgtta
      841 agaaaccagg gactaatgat tataggcctg tccaggatct gagagaagtc aacaagcggg
      901 tggaggacat ccaccccacc gtgcccaacc cttacaacct cttgagcggg ctcccaccgt
      961 cccaccagtg gtacactgtg cttgatttaa aggatgcctt tttctgcctg agactccacc
     1021 ccaccagtca gcctctcttc gcctttgagt ggagagatcc agagatggga atctcaggac
     1081 aattgacctg gaccagactc ccacagggtt tcaaaaacag tcccaccctg tttaatgagg
     1141 cactgcacag agacctagca gacttccgga tccagcaccc agacttgatc ctgctacagt
     1201 acgtggatga cttactgctg gccgccactt ctgagctaga ctgccaacaa ggtactcggg
     1261 ccctgttaca aaccctaggg aacctcgggt atcgggcctc ggccaagaaa gcccaaattt
     1321 gccagaaaca ggtcaagtat ctggggtatc ttctaaaaga gggtcagaga tggctgactg
     1381 aggccagaaa agagactgtg atggggcagc ctactcccaa gacccctcga caactaaggg
     1441 agttcctagg gaaggcaggc ttctgtcgcc tcttcatccc tgggtttgca gaaatggcag
     1501 cccccctgta ccctctcacc aaaccgggga ctctgtttaa ttggggccca gaccaacaaa
     1561 aggcctatca agaaatcaag caagctcttc taactgcccc agccctgggg ttgccagatt
     1621 tgactaagcc ctttgaactc tttgtcgacg agaagcaggg ctacgccaaa ggtgtcctaa
     1681 cgcaaaaact gggaccttgg cgtcggccgg tggcctacct gtccaaaaag ctagacccag
     1741 tagcagctgg gtggccccct tgcctacgga tggtagcagc cattgccgta ctgacaaagg
     1801 atgcaggcaa gctaaccatg ggacagccac tagtcattct ggccccccat gcagtagagg
     1861 cactagtcaa acaacccccc gaccgctggc tttccaacgc ccggatgact cactatcagg
     1921 ccttgctttt ggacacggac cgggtccagt tcggaccggt ggtagccctg aacccggcta
     1981 cgctgctccc actgcctgag gaagggctgc aacacaactg ccttgatatc ctggccgaag
     2041 cccacggaac ccgacccgac ctaacggacc agccgctccc agacgccgac cacacctggt
     2101 acacggatgg aagcagtctc ttacaagagg gacagcgtaa ggcgggagct gcggtgacca
     2161 ccgagaccga ggtaatctgg gctaaagccc tgccagccgg gacatccgct cagcgggctg
     2221 aactgatagc actcacccag gccctaaaga tggcagaagg taagaagcta aatgtttata
     2281 ctgatagccg ttatgctttt gctactgccc atatccatgg agaaatatac agaaggcgtg
     2341 ggtggctcac atcagaaggc aaagagatca aaaataaaga cgagatcttg gccctactaa
     2401 aagccctctt tctgcccaaa agacttagca taatccattg tccaggacat caaaagggac
     2461 acagcgccga ggctagaggc aaccggatgg ctgaccaagc ggcccgaaag gcagccatca
     2521 cagagactcc agacacctct accctcctca tagaaaattc atcaccctct ggcggctcaa
     2581 aaagaaccgc cgacggcagc gaattcgagc ccaagaagaa gaggaaagtc ggaagcacgt
     2641 cttcatcgga tcccgggccc gtcgactgca gaggcctgca tgcaagcttg gtgtaatcat
     2701 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     2761 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     2821 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     2881 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     2941 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     3001 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     3061 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     3121 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     3181 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     3241 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     3301 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     3361 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     3421 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     3481 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     3541 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     3601 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     3661 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     3721 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     3781 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaagc ccaatctgaa
     3841 taatgttaca accaattaac caattctgat tagaaaaact catcgagcat caaatgaaac
     3901 tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat
     3961 gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg
     4021 attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta
     4081 tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc
     4141 atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca
     4201 tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg
     4261 ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca
     4321 tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc tgtttttccg
     4381 gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc
     4441 ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg
     4501 gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaag
     4561 cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa
     4621 tcagcatcca tgttggaatt taatcgcggc ctcgacgttt cccgttgaat atggctcata
     4681 acaccccttg tattactgtt tatgtaagca gacagtttta ttgttcatga tgatatattt
     4741 ttatcttgtg caatgtaaca tcagagattt tgagacacgg gccagagctg ca
//
