LOCUS      pADH139       3094 bp    DNA     circular SYN 14-APR-2025
DEFINITION  NAT-marked ""empty"" gRNA construct for use in gRNA expression 
            cassette stitching PCR; use with pADH110 to generate target-specific
            part 2 of 2 for C.mal LEUpOUT CRISPR system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3094)
  AUTHORS   Nguyen N, Quail MMF, Hernday AD
  TITLE     An Efficient, Rapid, and Recyclable System for CRISPR-Mediated 
            Genome Editing in Candida albicans.
  JOURNAL   mSphere. 2017 Apr 26;2(2):e00149-17. doi: 
            10.1128/mSphereDirect.00149-17. eCollection 2017 Mar-Apr.
  PUBMED    28497115
REFERENCE   2  (bases 1 to 3094)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            //gt12365.cn
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1128/mSphereDirect.00149-17"; journalName: "mSphere"; date: 
            "2017-04-26- 26"; volume: "2"; issue: "2"; pages: "e00149-17"
COMMENT     Sequence Label: pADH139
FEATURES             Location/Qualifiers
     source          1..3094
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     138..156
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_RNA        190..265
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(join(1086..1092,1093..1110,1111..1116))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1131..1152
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1269..1286)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1440..2028)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1520..1539)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2199..2990,2991..3059))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATIDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2822..2841
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(3060..3094,1..70))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
       61 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
      121 tgacattaac ctataaaaat aggcgtatca cgaggccctt tgacggcacg gccacgcgtt
      181 taaaccgccg ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact
      241 tgaaaaagtg gcaccgagtc ggtgcttttt ttctcgagtt tttttatcga gttgagacca
      301 gaagtcgtta aaggtacaaa cttgatcatt gttcgtgaat tagttggtgg tatttatttc
      361 ggtgatcgtg aagaacaaga agaaagtgaa gataaacaaa ccgcatggga tactgaaaaa
      421 tacaccgtcg atgaagtcac cagaatcacc cgtatggctg ccttcatggc tttacaacat
      481 aacccaccat taccaatctg gtcattggat aaggctaatg ttttagcttc ttctagatta
      541 tggagaagaa ctgtcgataa agtgatttct gaagagttcc caactttgtc tgttcaacat
      601 caattgattg actccgccgc catgatttta attcaaaacc caaccaaatt gaatggtata
      661 atcatcactt ctaacatgtt tggtgatatc atttccgatg aagcttcagt tattccaggt
      721 tccttgggtt tattaccatc tgcatctttg gcttcattgc cagataccaa caccgctttt
      781 ggtctttatg aaccatgtca cggttctgct cctgatttac cagccaataa agttaaccca
      841 attgctacta ttttatctgc tgcttctatg ttgagattat ctttggattg tgtgaaagaa
      901 gctgaagctt tagaagaagc cgtgaaacaa gtcttggata aaggtatcag aactgcagat
      961 ttaagaggta ccagttcagc tactgaagtt ggtgacgcga ttgtcgaagc tgttactaaa
     1021 atcttaaaag aaaaagctta acggtgttta aaccccagcg cctggcgggc gctcacaatt
     1081 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     1141 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     1201 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     1261 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     1321 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     1381 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     1441 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     1501 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     1561 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     1621 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     1681 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     1741 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     1801 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     1861 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     1921 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     1981 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     2041 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     2101 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     2161 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt gatcagtgag
     2221 gcacctatct cagcgatctg tctatttcgt tcatcaatag ttgcctgact ccccgtcgtg
     2281 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     2341 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     2401 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     2461 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     2521 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     2581 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     2641 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     2701 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     2761 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     2821 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     2881 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     2941 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     3001 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     3061 ctcttccttt ttcaatatta ttgaagcatt tatc
//