LOCUS       pADH147       2878 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 HIS1-FLP CRISPR system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2878)
  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 2878)
  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: pADH147
FEATURES             Location/Qualifiers
     source          1..2878
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     111..130
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(349..453)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     521..539
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_RNA        573..648
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(675..708)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     promoter        complement(join(1253..1259,1260..1277,1278..1283))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1298..1319
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1436..1453)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1607..2195)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1687..1706)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2366..2878,1..279,280..348))
                     /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"
ORIGIN
        1 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
       61 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
      121 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
      181 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
      241 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
      301 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
      361 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
      421 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
      481 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
      541 ctttgacggc acggccacgc gtttaaaccg ccgttttaga gctagaaata gcaagttaaa
      601 ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt tttttctcga
      661 gtttttttat cgaggaagtt cctatacttt ctagagaata ggaacttccc accttgatgt
      721 acacaccttt aaagtcaata ggcattctcg tttgtaattc cttgacacac atatccatta
      781 actttaaacc atcttcgacc gtcatgtcct ttctataatg tttatctaac aaagatgttg
      841 tgtaaaatgc agcgtagcca tgagcaccat aagggagttc aacttgtgta cccaaataat
      901 ctatccaatt caacgatggc tcatttttct ttacatcata tcctccaatt aaacagttca
      961 cctggtacgg ttttctagat ctaagggaac tggccaattg gtttctcaca aacgaggctg
     1021 ttgcctttgg agataactca atgtcatttt ctctcatcga gtacaattga atgtttgctt
     1081 gaacatactc ggcaaactga accgtatcac cagcttcacc agtgtatgct attaagttgt
     1141 gtgcatttaa atgtcttgtt ttatcatctg tatcctttaa aattgaaata cctcttgtgg
     1201 cagctttacg gtgtttaaac cccagcgcct ggcgggcgct cacaattcca cacaacatac
     1261 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     1321 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     1381 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     1441 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     1501 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     1561 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     1621 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     1681 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     1741 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     1801 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     1861 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     1921 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     1981 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     2041 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     2101 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     2161 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     2221 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     2281 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     2341 tatatgagta aacttggtct gacagttacc aatgcttgat cagtgaggca cctatctcag
     2401 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     2461 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     2521 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     2581 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     2641 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     2701 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     2761 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     2821 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttac
//