LOCUS       sequence-431531-        4834 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fragmid fragment: (C' terminus) for prime editing; using PEmax.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4834)
  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 4834)
  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: AA184
FEATURES             Location/Qualifiers
     source          1..4834
                     /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             495..515
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2616..2636
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2646..2672
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     primer_bind     complement(2744..2760)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2744..2760)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2757..2779)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2768..2784
                     /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(2792..2798,2799..2816,2817..2822))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2837..2858
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2975..2992)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3146..3734)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3226..3245)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3912..4721)
                     /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     4635..4654
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4713..4732
                     /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 tccggaagct ctggtggcag caagcggacc gccgacggct
      481 ctgaattcga gagccctaag aagaaaagaa aggtgagcgg aggctctagc ggcggaagca
      541 ccctgaacat tgaggacgag tatagactgc atgaaacaag caaggaaccc gacgtgtccc
      601 tgggctccac ctggctgtcc gactttcccc aggcctgggc cgagacagga ggaatgggcc
      661 tggccgtgcg gcaggcaccc ctgatcatcc ctctgaaggc cacctctaca cccgtgagca
      721 tcaagcagta ccctatgtct caggaggcca gactgggcat caagcctcac atccagaggc
      781 tgctggacca gggcatcctg gtgccatgcc agagcccctg gaacacacca ctgctgcccg
      841 tgaagaagcc aggcaccaat gactatagac ccgtgcagga tctgagagag gtgaacaaga
      901 gggtggagga tatccacccc accgtgccca acccttacaa tctgctgtcc ggcctgcccc
      961 cttctcacca gtggtataca gtgctggacc tgaaggatgc cttcttttgt ctgagactgc
     1021 accctaccag ccagccactg ttcgcctttg agtggaggga ccctgagatg ggcatctctg
     1081 gccagctgac ctggacacgc ctgcctcagg gcttcaagaa tagcccaaca ctgtttaacg
     1141 aggccctgca ccgcgacctg gcagatttcc ggattcagca cccagatctg atcctgctgc
     1201 agtacgtgga cgatctgctg ctggccgcca ccagcgagct ggattgccag cagggaacac
     1261 gcgccctgct gcagaccctg ggaaacctgg gatatagggc atccgccaag aaggcccaga
     1321 tctgtcagaa gcaggtgaag tacctgggct atctgctgaa ggagggccag agatggctga
     1381 cagaggccag gaaggagaca gtgatgggcc agccaacacc caagacccca agacagctga
     1441 gggagttcct gggcaaagca ggattttgca ggctgttcat cccaggattc gcagagatgg
     1501 cagcacctct gtacccactg accaagccgg gcaccctgtt taattggggc cctgaccagc
     1561 agaaggccta tcaggagatc aagcaggccc tgctgacagc accagccctg ggcctgccag
     1621 acctgaccaa gcctttcgag ctgtttgtgg atgagaagca gggctacgcc aagggcgtgc
     1681 tgacccagaa gctgggacca tggaggcggc ccgtggccta tctgtccaag aagctggacc
     1741 cagtggcagc aggatggcca ccatgcctga ggatggtggc agcaatcgcc gtgctgacaa
     1801 aggatgccgg caagctgacc atgggacagc cactggtcat cctggcacca cacgcagtgg
     1861 aggccctggt gaagcagcct ccagatcgct ggctgtctaa cgcccggatg acacactacc
     1921 aggccctgct gctggacacc gatcgcgtgc agtttggccc tgtggtggcc ctgaatccag
     1981 ccaccctgct gcctctgcca gaggagggcc tgcagcacaa ctgtctggac atcctggcag
     2041 aggcacacgg aacaaggcca gacctgaccg atcagcccct gcctgacgcc gatcacacat
     2101 ggtataccga tggaagctcc ctgctgcagg agggccagag gaaggcagga gcagcagtga
     2161 ccacagagac agaagtgatc tgggccaagg ccctgccagc aggcacatcc gcccagcggg
     2221 ccgagctgat cgccctgacc caggccctga agatggccga gggcaagaag ctgaacgtgt
     2281 acacagactc cagatatgcc ttcgccaccg cacacatcca cggagagatc tacaggcgcc
     2341 ggggctggct gacctctgag ggcaaggaga tcaagaacaa ggatgagatc ctggccctgc
     2401 tgaaggccct gtttctgccc aagcggctga gcatcatcca ctgtcctgga caccagaagg
     2461 gacactccgc cgaggcaagg ggcaatcgga tggccgacca ggccgccaga aaggctgcta
     2521 ttactgaaac tcccgacact tccactctgc tgattgaaaa ctcctcccct tctggcggct
     2581 caaaaagaac cgccgacggc agcgaattcg agtctcccaa gaagaagagg aaagtcggat
     2641 ccggccctgc cgctaagaga gtgaagctgg acggaagcac gtcttcatcg gatcccgggc
     2701 ccgtcgactg cagaggcctg catgcaagct tggtgtaatc atggtcatag ctgtttcctg
     2761 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     2821 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     2881 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     2941 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3001 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3061 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3121 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3181 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3241 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3301 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3361 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3421 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3481 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3541 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     3601 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3661 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3721 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3781 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3841 ttttaaatta aaaatgaagt tttaaatcaa gcccaatctg aataatgtta caaccaatta
     3901 accaattctg attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca
     3961 ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg
     4021 aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca
     4081 tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca
     4141 tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt
     4201 tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc
     4261 attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa
     4321 acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct
     4381 gaatcaggat attcttctaa tacctggaat gctgtttttc cggggatcgc agtggtgagt
     4441 aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc
     4501 gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca
     4561 tgtttcagaa acaactctgg cgcatcgggc ttcccataca agcgatagat tgtcgcacct
     4621 gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa
     4681 tttaatcgcg gcctcgacgt ttcccgttga atatggctca taacacccct tgtattactg
     4741 tttatgtaag cagacagttt tattgttcat gatgatatat ttttatcttg tgcaatgtaa
     4801 catcagagat tttgagacac gggccagagc tgca
//
