LOCUS       sequence-381843-        2970 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mutant I-F CRISPR Leader and a CRISPR locus with two consensus I-F 
            repeats, for integration assays..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2970)
  AUTHORS   Santiago-Frangos A, Henriques WS, Wiegand T, Gauvin CC, Buyukyoruk 
            M, Graham AB, Wilkinson RA, Triem L, Neselu K, Eng ET, Lander GC, 
            Wiedenheft B
  TITLE     Structure reveals why genome folding is necessary for site-specific 
            integration of foreign DNA into CRISPR arrays.
  JOURNAL   Nat Struct Mol Biol. 2023 Nov;30(11):1675-1685. doi: 
            10.1038/s41594-023-01097-2. Epub 2023 Sep 14.
  PUBMED    37710013
REFERENCE   2  (bases 1 to 2970)
  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"; doi: 
            "10.1038/s41594-023-01097-2"; journalName: "Nat Struct Mol Biol"; 
            date: "2023-11"; volume: "30"; issue: "11"; pages: "1675-1685"
COMMENT     Sequence Label: pX1-IFl_Con-IFr
FEATURES             Location/Qualifiers
     source          1..2970
                     /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"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(749..765)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(749..765)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(762..784)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    773..789
                     /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(797..803,804..821,822..827))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    842..863
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(980..997)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1151..1739)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1231..1250)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1910..2701,2702..2770))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2533..2552
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2771..2875)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2943..2961
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     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 ccagtgaatt cgagctcggt acctcgcgaa
      421 tgcatctaga tccatggacc cttttttcgg acttaaaaaa cccttataaa tcagcaagtt
      481 acgagacctc gaaaaaaaag ggtttctggc gggaaaaact caaaaaacgc ttcgaaatca
      541 accggttata ggttttcgga gctagttcac tgccgtgtag gcagctaaga aaatcagccg
      601 gacgttgtag tagtcgagcg cggtgttcac tgccgtgtag gcagctaaga aagccgggtg
      661 gtgacttctg gacggcgttc gtcaaggatc catcggatcc cgggcccgtc gactgcagag
      721 gcctgcatgc aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
      781 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
      841 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
      901 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
      961 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     1021 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     1081 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     1141 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     1201 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     1261 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     1321 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     1381 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     1441 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     1501 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     1561 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     1621 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     1681 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     1741 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     1801 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     1861 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     1921 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     1981 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     2041 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     2101 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     2161 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     2221 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     2281 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     2341 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     2401 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     2461 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     2521 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     2581 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     2641 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     2701 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     2761 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     2821 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     2881 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     2941 aaaataggcg tatcacgagg ccctttcgtc
//
