LOCUS       sequence-431536-        4253 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fragmid fragment: (Cas protein) firefly luciferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4253)
  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 4253)
  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: AA201
FEATURES             Location/Qualifiers
     source          1..4253
                     /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             445..2091
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="EDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAH
                     IEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPA
                     NDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSM
                     YTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHAR
                     DPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKI
                     QSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYG
                     LTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMSG
                     YVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILL
                     QHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVF
                     VDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(508..526)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     primer_bind     complement(2163..2179)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2163..2179)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2176..2198)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2187..2203
                     /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(2211..2217,2218..2235,2236..2241))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2256..2277
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2394..2411)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2565..3153)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2645..2664)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3331..4140)
                     /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     4054..4073
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4132..4151
                     /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 tgaagactgg cagcgaggac gccaaaaaca taaagaaagg cccggcgcca
      481 ttctatccgc tggaagatgg aaccgctgga gagcaactgc ataaggctat gaagagatac
      541 gccctggttc ctggaacaat tgcttttaca gatgcacata tcgaggtgga catcacttac
      601 gctgagtact tcgaaatgtc cgttcggttg gcagaagcta tgaaacgata tgggctgaat
      661 acaaatcaca gaatcgtcgt atgcagtgaa aactctcttc aattctttat gccggtgttg
      721 ggcgcgttat ttatcggagt tgcagttgcg cccgcgaacg acatttataa tgaacgtgaa
      781 ttgctcaaca gtatgggcat ttcgcagcct accgtggtgt tcgtttccaa aaaggggttg
      841 caaaaaattt tgaacgtgca aaaaaagctc ccaatcatcc aaaaaattat tatcatggat
      901 tctaaaacgg attaccaggg atttcagtcg atgtacacgt tcgtcacatc tcatctacct
      961 cccggtttta atgaatacga ttttgtgcca gagtccttcg atagggacaa gacaattgca
     1021 ctgatcatga actcctctgg atctactggt ctgcctaaag gtgtcgctct gcctcataga
     1081 actgcctgcg tgagattctc gcatgccaga gatcctattt ttggcaatca aatcattccg
     1141 gatactgcga ttttaagtgt tgttccattc catcacggtt ttggaatgtt tactacactc
     1201 ggatatttga tatgtggatt tcgagtcgtc ttaatgtata gatttgaaga agagctgttt
     1261 ctgaggagcc ttcaggatta caagattcaa agtgcgctgc tggtgccaac cctattctcc
     1321 ttcttcgcca aaagcactct gattgacaaa tacgatttat ctaatttaca cgaaattgct
     1381 tctggtggcg ctcccctctc taaggaagtc ggggaagcgg ttgccaagag gttccatctg
     1441 ccaggtatca ggcaaggata tgggctcact gagactacat cagctattct gattacaccc
     1501 gagggggatg ataaaccggg cgcggtcggt aaagttgttc cattttttga agcgaaggtt
     1561 gtggatctgg ataccgggaa aacgctgggc gttaatcaaa gaggcgaact gtgtgtgaga
     1621 ggtcctatga ttatgtccgg ttatgtaaac aatccggaag cgaccaacgc cttgattgac
     1681 aaggatggat ggctacattc tggagacata gcttactggg acgaggacga acacttcttc
     1741 atcgttgacc gcctgaagtc tctgattaag tacaaaggct atcaggtggc tcccgctgaa
     1801 ttggaatcca tcttgctcca acaccccaac atcttcgacg caggtgtcgc aggccttccc
     1861 gacgatgacg ccggtgaact tcccgccgcc gttgttgttt tggagcacgg aaagacgatg
     1921 acggaaaaag agatcgtgga ttacgtcgcc agtcaagtaa caaccgcgaa aaagttgcgc
     1981 ggaggagttg tgtttgtgga cgaagtaccg aaaggtctta ccggaaaact cgacgcaaga
     2041 aaaatcagag agatcctcat aaaggccaag aagggcggaa agatcgccgt ggggtccaag
     2101 tcttcatcgg atcccgggcc cgtcgactgc agaggcctgc atgcaagctt ggtgtaatca
     2161 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     2221 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     2281 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     2341 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     2401 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     2461 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     2521 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2581 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2641 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2701 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2761 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2821 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2881 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     2941 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     3001 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     3061 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     3121 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     3181 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     3241 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaag cccaatctga
     3301 ataatgttac aaccaattaa ccaattctga ttagaaaaac tcatcgagca tcaaatgaaa
     3361 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     3421 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     3481 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     3541 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg
     3601 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     3661 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct
     3721 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     3781 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgtttttcc
     3841 ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     3901 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     3961 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     4021 gcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     4081 atcagcatcc atgttggaat ttaatcgcgg cctcgacgtt tcccgttgaa tatggctcat
     4141 aacacccctt gtattactgt ttatgtaagc agacagtttt attgttcatg atgatatatt
     4201 tttatcttgt gcaatgtaac atcagagatt ttgagacacg ggccagagct gca
//
