LOCUS       sequence-431457-        2756 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fragmid fragment: (N' terminus) attenuate expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2756)
  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 2756)
  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: AA371
FEATURES             Location/Qualifiers
     source          1..2756
                     /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             536..562
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             569..595
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     primer_bind     complement(666..682)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(666..682)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(679..701)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    690..706
                     /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(714..720,721..738,739..744))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    759..780
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(897..914)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1068..1656)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1148..1167)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1834..2643)
                     /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     2557..2576
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2635..2654
                     /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 tgaagacgta ggagaattcc aacatggtga gcaagggcga ggagctgttc
      481 accggggtgg tgcccatcct ggtcgagctg gacggctcta gataataaca tggccctgcc
      541 tccactggaa cggctgaccc tggggtcgct gccacccctg gagcgcctga ccctgggcag
      601 cagtcttcat cggatcccgg gcccgtcgac tgcagaggcc tgcatgcaag cttggtgtaa
      661 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
      721 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
      781 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
      841 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
      901 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
      961 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     1021 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     1081 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     1141 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     1201 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     1261 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     1321 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     1381 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     1441 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     1501 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     1561 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     1621 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     1681 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     1741 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aagcccaatc
     1801 tgaataatgt tacaaccaat taaccaattc tgattagaaa aactcatcga gcatcaaatg
     1861 aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa gccgtttctg
     1921 taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct ggtatcggtc
     1981 tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt caaaaataag
     2041 gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg gcaaaagttt
     2101 atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat caaaatcact
     2161 cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa atacgcgatc
     2221 gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga acactgccag
     2281 cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga atgctgtttt
     2341 tccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat
     2401 ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc
     2461 attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata
     2521 caagcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata
     2581 taaatcagca tccatgttgg aatttaatcg cggcctcgac gtttcccgtt gaatatggct
     2641 cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc atgatgatat
     2701 atttttatct tgtgcaatgt aacatcagag attttgagac acgggccaga gctgca
//
