LOCUS       pADH119        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.alb 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: pADH119
FEATURES             Location/Qualifiers
     source          1..3094
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     173..191
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_RNA        225..300
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(join(1121..1127,1128..1145,1146..1151))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1166..1187
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1304..1321)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1475..2063)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1555..1574)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2234..3025,3026..3094))
                     /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     2857..2876
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
      181 tatcacgagg ccctttgacg gcacggccac gcgtttaaac cgccgtttta gagctagaaa
      241 tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc gagtcggtgc
      301 tttttttctc gagttttttt atcgagttaa aagctgaagt ggtcaaagga accaatttaa
      361 ttattgttcg tgaattagtt ggtggtatat attttggtga acgtcaagaa caagaagaaa
      421 gtgaagacaa aaaaacagct tgggatacag aaaagtatac tgttgacgaa gttactcgta
      481 tcactcgtat ggcagcattt atggctttgc agcacaatcc accattgcca atatggtctt
      541 tggacaaagc taatgttttg gcatcgtcga gattgtggag aaagaccgtt gataaagtga
      601 tttctgaaga atttccagct ttgtctgtgc agcatcaatt gattgattcg gctgcaatga
      661 ttttgatcca aaacccaacc aaattgaacg gtatcatcat tacatctaac atgtttggtg
      721 atatcatctc agatgaggca tccgttatcc caggatcttt gggtttattg ccatctgctt
      781 cattggcttc attgccagat accaacactg cttttggtct ttatgagcct tgccatgggt
      841 ctgctccaga tttaccagcc aacaaagtga atccaattgc tacaatcttg tctgccgctt
      901 ccatgttgag attatccttg gattgtgtta aggaagctga agcattggaa gaagctgtca
      961 aacaggtttt ggatagcggt atcagaacag cagatttaag aggtactagc tctactacag
     1021 aagtcggtga cgcaatagcc gaaacagtca ctaaaatttt aaaacaaaat acataacggt
     1081 gtttaaaccc cagcgcctgg cgggcgctca caattccaca caacatacga gccggaagca
     1141 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     1201 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     1261 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     1321 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     1381 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     1441 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     1501 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     1561 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     1621 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     1681 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     1741 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     1801 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     1861 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     1921 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     1981 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     2041 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     2101 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     2161 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     2221 cttggtctga cagttaccaa tgcttgatca gtgaggcacc tatctcagcg atctgtctat
     2281 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     2341 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     2401 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     2461 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     2521 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     2581 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     2641 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     2701 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     2761 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     2821 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     2881 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     2941 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     3001 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     3061 gaataagggc gacacggaaa tgttgaatac tcat
//