LOCUS       sequence-301640-        7044 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  P. aeruginosa PA14 CRISPR2 locus, with ""optimized"" IHFprox site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7044)
  AUTHORS   Santiago-Frangos A, Buyukyoruk M, Wiegand T, Krishna P, Wiedenheft B
  TITLE     Distribution and phasing of sequence motifs that facilitate CRISPR 
            adaptation.
  JOURNAL   Curr Biol. 2021 Aug 23;31(16):3515-3524.e6. doi: 
            10.1016/j.cub.2021.05.068. Epub 2021 Jun 25.
  PUBMED    34174210
REFERENCE   2  (bases 1 to 7044)
  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: 
            "10.1016/j.cub.2021.05.068"; journalName: "Curr Biol"; date: 
            "2021-08-23- 23"; volume: "31"; issue: "16"; pages: "3515-3524.e6"
COMMENT     Sequence Label: pCRISPR2_IHF_Opt
FEATURES             Location/Qualifiers
     source          1..7044
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             37..59
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(1994..2010)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2003..2025)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(2171..3004)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pRO1600 from Pseudomonas aeruginosa "
                     /label=pRO1600 Rep
                     /translation="MASPPMVYKSNALVEAAYRLSVQEQRIVLACISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMDSTHAIRLYELLMQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKAGKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     rep_origin      3018..3369
                     /label=pRO1600 oriV
                     /note="broad-host-range origin of replication from 
                     Pseudomonas aeruginosa plasmid pRO1600; requires the 
                     pRO1600 Rep protein for replication (West et al., 1994)"
     promoter        join(3410..3415,3416..3432,3433..3438)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             3608..4141
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLWSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     rep_origin      4942..5530
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5431..5450
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     oriT            5600..5708
                     /note="incP origin of transfer"
     CDS             complement(5865..6743)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        join(6770..7044,1..10)
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     6977..6996
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 gtttctccat acccgttttt ttgggctaga aataattttg tttaacttta agaaggagat
       61 atacataccc atggatgcct tgtccgaaat cgaggcgggc cacccgagcc tgtagcgccg
      121 ctccctggcg cgacgccccc ggcctggccg ggggctttgc ggcatcgccc atcaaaagac
      181 ctttcgcgcc cgaacggcac ggttgatcgc cgtcccggtc ctcgcgaaac ggccgccaat
      241 tgcccgaagc ttccgaccct tttttcggac gatttcttac gcccttataa atcagcaagt
      301 tacgagacct cgaaaaaaga gggtttctgg cgggaaaaac tcaaaaaacg cttcgaaatc
      361 aaccggttat aggttttcgg agctagttca ctgccgtgta ggcagctaag aaaatcagcc
      421 ggacgttgta gtagtcgagc gcggtgttca ctgccgtgta ggcagctaag aaagccgggt
      481 ggtgacttct ggacggcgtt cgtcaagttc actgccgtgt aggcagctaa gaaagacctg
      541 gttttggccg acacagtggc tgcgacgttc actgccgtgt aggcagctaa gaaacggtcc
      601 tgccagtact gcggcaggat ctggtcgttc actgccgtgt aggcagctaa gaaaagcctg
      661 gcgcccttcg agtccatcca gcccgggttc actgccgtgt aggcagctaa gaaacatcca
      721 attcgccgat ccggcgcaag cgcccggttc actgccgtgt aggcagctaa gaaaacgaag
      781 cagatcgccg aatgcggcgt cagcgcgttc actgccgtgt aggcagctaa gaaagtgggt
      841 gcaaatcgcg attctggtcg cgtcgtgttc actgccgtgt aggcagctaa gaaatgaata
      901 tcagcctgca aagcgtcgtc gtagtagttc actgccgtgt aggcagctaa gaaaaagccg
      961 ggccgatgtt cgagcctggc tgcgtggttc actgccgtgt aggcagctaa gaaatggaca
     1021 atgcgcagcc caggcgctgg atttacgttc actgccgtgt aggcagctaa gaaaatgcga
     1081 ggctgatcga ggtgcgcggt tctgttgttc actgccgtgt aggcagctaa gaaagtgcgc
     1141 ggatggctgg tatctcgagc acgattgttc actgccgtgt aggcagctaa gaaaaggcaa
     1201 acaacggaaa gatcaccacc gaaggcgttc actgccgtgt aggcagctaa gaaaaacgaa
     1261 acaggcccag cacagccgct tgataagttc actgccgtgt aggcagctaa gaaaacaaga
     1321 cgctcatcga cggcagcatc gagacggttc actgccgtgt aggcagctaa gaaaaaggtg
     1381 acgatgcaca gctgttgcgc gcggttgttc actgccgtgt aggcagctaa gaaaatcctc
     1441 gaacagcccg ggttcggtcg cctccagttc actgccgtgt aggcagctaa gaaaaggagt
     1501 ttgcggcccc gctccctgcg gctctcgttc actgccgtgt aggcagctaa gaaattcacg
     1561 gcgggcttga tgtccgcgtc tacctggttc actgccgtgt aggcagctaa gaaattgccg
     1621 agtacgatgc ctgatacatg aatccagttc actgccgtgt aggcagctaa gaaactcgaa
     1681 cccacctcgg ccacaacagc cgccgggttc gctgccgtct aggcagaacc accctcccca
     1741 tcccactacc aaacatccga atataaagtt cctaccccgc ccgccagcct cgcccgtcca
     1801 cgacaatgtg cctcgcttgg aagctcacgc tcctcacaca gacgaaaacg gggtagcggc
     1861 aatcggccat atccgctaaa cagttgcctt ggcgcagaat tcgatagatc cgatagggac
     1921 aggccacggt caacatggac gacatttctc ccagcgaact gaagacgatc cttcactcgc
     1981 atgcaagctt ggcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     2041 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     2101 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa aggcaggccg ggccgtggtg
     2161 gccacggcct ctaggccaga tccagcggca tctgggttag tcgagcgcgg gccgcttccc
     2221 atgtctcacc agggcgagcc tgtttcgcga tctcagcatc tgaaatcttc ccggccttgc
     2281 gcttcgctgg ggccttaccc accgccttgg cgggcttctt cggtccaaaa ctgaacaaca
     2341 gatgtgtgac cttgcgcccg gtctttcgct gcgcccactc cacctgtagc gggctgtgct
     2401 cgttgatctg cgtcacggct ggatcaagca ctcgcaactt gaagtccttg atcgagggat
     2461 accggccttc cagttgaaac cactttcgca gctggtcaat ttctatttcg cgctggccga
     2521 tgctgtccca ttgcatgagc agctcgtaaa gcctgatcgc gtgggtgctg tccatcttgg
     2581 ccacgtcagc caaggcgtat ttggtgaact gtttggtgag ttccgtcagg tacggcagca
     2641 tgtctttggt gaacctgagt tctacacggc cctcaccctc ccggtagatg attgtttgca
     2701 cccagccggt aatcatcaca ctcggtcttt tccccttgcc attgggctct tgggttaacc
     2761 ggacttcccg ccgtttcagg cgcagggccg cttctttgag ctggttgtag gaagattcga
     2821 tagggacacc cgccatcgtc gctatgtcct ccgccgtcac tgaatacatc acttcatcgg
     2881 tgacaggctc gctcctcttc acctggctaa tacaggccag aacgatccgc tgttcctgaa
     2941 cactgaggcg atacgcggcc tcgaccaggg cattgctttt gtaaaccatt gggggtgagg
     3001 ccacgttcga cattccttgt gtataagggg acactgtatc tgcgtcccac aatacaacaa
     3061 atccgtccct ttacaacaac aaatccgtcc cttcttaaca acaaatccgt cccttaatgg
     3121 caacaaatcc gtcccttttt aaactctaca ggccacggat tacgtggcct gtagacgtcc
     3181 taaaaggttt aaaagggaaa aggaagaaaa gggtggaaac gcaaaaaacg caccactacg
     3241 tggccccgtt ggggccgcat ttgtgcccct gaaggggcgg gggaggcgtc tgggcaatcc
     3301 ccgttttacc agtcccctat cgccgcctga gagggcgcag gaagcgagta atcagggtat
     3361 cgaggcggat tcacccttgg cgtccaacca gcggcaccag cggcgcgaat tgacataagc
     3421 ctgttcggtt cgtaaactgt aatgcaagta gcgtatgcgc tcacgcaact ggtccagaac
     3481 cttgaccgaa cgcagcggtg gtaacggcgc agtggcggtt ttcatggctt gttatgactg
     3541 tttttttgta cagtctatgc ctcgggcatc caagcagcaa gcgcgttacg ccgtgggtcg
     3601 atgtttgatg ttatggagca gcaacgatgt tacgcagcag ggcagtcgcc ctaaaacaaa
     3661 gttaggtggc tcaagtatgg gcatcattcg cacatgtagg ctcggccctg accaagtcaa
     3721 atccatgcgg gctgctcttg atcttttcgg tcgtgagttc ggagacgtag ccacctactc
     3781 ccaacatcag ccggactccg attacctcgg gaacttgctc cgtagtaaga cattcatcgc
     3841 gcttgctgcc ttcgaccaag aagcggttgt tggcgctctc gcggcttacg ttctgcccaa
     3901 gtttgagcag ccgcgtagtg agatctatat ctatgatctc gcagtctccg gcgagcaccg
     3961 gaggcagggc attgccaccg cgctcatcaa tctcctcaag catgaggcca acgcgcttgg
     4021 tgcttatgtg atctacgtgc aagcagatta cggtgacgat cccgcagtgg ctctctatac
     4081 aaagttgggc atacgggaag aagtgatgca ctttgatatc gacccaagta ccgccaccta
     4141 acaattcgtt caagccgaga tcggcttccc ggccgcggag ttgttcggta aattgtcaca
     4201 acgccgcggc caattcgact caccagtcac agaaaagcat cttacggatg gcatgacagt
     4261 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     4321 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     4381 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     4441 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     4501 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     4561 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     4621 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     4681 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     4741 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     4801 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     4861 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     4921 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     4981 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5041 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     5101 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     5161 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     5221 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     5281 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     5341 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     5401 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     5461 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     5521 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     5581 tgctcacaag ctagcttatc ggccagcctc gcagagcagg attcccgttg agcaccgcca
     5641 ggtgcgaata agggacagtg aagaaggaac acccgctcgc gggtgggcct acttcaccta
     5701 tcctgcccgg ctgacgccgt tggatacacc aaggaaagtc tacacgaacc ctttggcaaa
     5761 atcctgtata tcgtgcgaaa aaggatggat ataccgaaaa aatcgctata atgaccccga
     5821 agcagggtta tgcagcggaa agtatacctt aaggaatccc caaattatga caacttgacg
     5881 gctacatcat tcactttttc ttcacaaccg gcacggaact cgctcgggct ggccccggtg
     5941 cattttttaa atacccgcga gaaatagagt tgatcgtcaa aaccaacatt gcgaccgacg
     6001 gtggcgatag gcatccgggt ggtgctcaaa agcagcttcg cctggctgat acgttggtcc
     6061 tcgcgccagc ttaagacgct aatccctaac tgctggcgga aaagatgtga cagacgcgac
     6121 ggcgacaagc aaacatgctg tgcgacgctg gcgatatcaa aattgctgtc tgccaggtga
     6181 tcgctgatgt actgacaagc ctcgcgtacc cgattatcca tcggtggatg gagcgactcg
     6241 ttaatcgctt ccatgcgccg cagtaacaat tgctcaagca gatttatcgc cagcagctcc
     6301 gaatagcgcc cttccccttg cccggcgtta atgatttgcc caaacaggtc gctgaaatgc
     6361 ggctggtgcg cttcatccgg gcgaaagaac cccgtattgg caaatattga cggccagtta
     6421 agccattcat gccagtaggc gcgcggacga aagtaaaccc actggtgata ccattcgcga
     6481 gcctccggat gacgaccgta gtgatgaatc tctcctggcg ggaacagcaa aatatcaccc
     6541 ggtcggcaaa caaattctcg tccctgattt ttcaccaccc cctgaccgcg aatggtgaga
     6601 ttgagaatat aacctttcat tcccagcggt cggtcgataa aaaaatcgag ataaccgttg
     6661 gcctcaatcg gcgttaaacc cgccaccaga tgggcattaa acgagtatcc cggcagcagg
     6721 ggatcatttt gcgcttcagc catacttttc atactcccgc cattcagaga agaaaccaat
     6781 tgtccatatt gcatcagaca ttgccgtcac tgcgtctttt actggctctt ctcgctaacc
     6841 aaaccggtaa ccccgcttat taaaagcatt ctgtaacaaa gcgggaccaa agccatgaca
     6901 aaaacgcgta acaaaagtgt ctataatcac ggcagaaaag tccacattga ttatttgcac
     6961 ggcgtcacac tttgctatgc catagcattt ttatccataa gattagcgga tcctacctga
     7021 cgctttttat cgcaactctc tact
//
