LOCUS       sequence-431416-        3018 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fragmid fragment: (guide cassette) guide expression; enables iBar 
            UMIs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3018)
  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 3018)
  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: AA107
FEATURES             Location/Qualifiers
     source          1..3018
                     /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"
     promoter        444..692
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     444..464
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     615..634
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(928..944)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(928..944)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(941..963)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    952..968
                     /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(976..982,983..1000,1001..1006))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1021..1042
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1159..1176)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1330..1918)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1410..1429)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2096..2905)
                     /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     2819..2838
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2897..2916
                     /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 tgaagacgtc agagagggcc tatttcccat gattccttca tatttgcata
      481 tacgatacaa ggctgttaga gagataatta gaattaattt gactgtaaac acaaagatat
      541 tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat
      601 tatgttttaa aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg
      661 ctttatatat cttgtggaaa ggacgaaaca ccggtaattt ctactcttgt agatggagac
      721 gcgttttttc tcgtctctta gcctggtctt gaaaaagtgg caccgagtcg gtgaattcgc
      781 tagcaagctt ggcgtaacta gatcttgaga caaatggcag tattcatcca caaaggtctt
      841 gaaaggagtg ggaattggac acgcgtcttc atcggatccc gggcccgtcg actgcagagg
      901 cctgcatgca agcttggtgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
      961 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     1021 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     1081 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     1141 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     1201 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     1261 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     1321 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     1381 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     1441 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     1501 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     1561 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     1621 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     1681 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     1741 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     1801 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     1861 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     1921 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     1981 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     2041 aagttttaaa tcaagcccaa tctgaataat gttacaacca attaaccaat tctgattaga
     2101 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     2161 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     2221 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     2281 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     2341 ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat
     2401 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     2461 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     2521 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     2581 ctaatacctg gaatgctgtt tttccgggga tcgcagtggt gagtaaccat gcatcatcag
     2641 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     2701 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     2761 ctggcgcatc gggcttccca tacaagcgat agattgtcgc acctgattgc ccgacattat
     2821 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctcg
     2881 acgtttcccg ttgaatatgg ctcataacac cccttgtatt actgtttatg taagcagaca
     2941 gttttattgt tcatgatgat atatttttat cttgtgcaat gtaacatcag agattttgag
     3001 acacgggcca gagctgca
//
