LOCUS       Exported                6227 bp ds-DNA    circular SYN 06-1-2016
DEFINITION  Drosophila expression vector in which a guide RNA (gRNA) is 
            expressed from the U6-2 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pCFD2-dU6-2gRNA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6227)
  AUTHORS   Port F, Chen HM, Lee T, Bullock SL.
  TITLE     Optimized CRISPR/Cas tools for efficient germline and somatic genome
            engineering in Drosophila.
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 2014;111:E2967-76.
  PUBMED    25002478
REFERENCE   2  (bases 1 to 6227)
  AUTHORS   Bullock Lab / Addgene #49409
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6227
                     /organism="synthetic DNA construct"
                     /lab_host="Drosophila melanogaster"
                     /mol_type="other DNA"
     promoter        7..406
                     /note="dU6-2 promoter"
                     /note="RNA polymerase III promoter for Drosophila U6-2 
                     snRNA (Das et al., 1987)"
     misc_RNA        425..500
                     /note="gRNA scaffold"
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      501..506
                     /note="polIII terminator"
                     /note="RNA polymerase III transcription terminator"
     rep_origin      complement(1252..1840)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2011..2871)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(2872..2976)
                     /gene="bla"
                     /note="AmpR promoter"
     rep_origin      complement(3002..3457)
                     /direction=LEFT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     CDS             complement(join(3832..3949,4026..4119,4211..4817,4872..5005,
                     5066..5226,5300..5325))
                     /codon_start=1
                     /product="tryptophan oxygenase"
                     /note="vermilion"
                     /note="selectable marker, required to synthesize the brown 
                     eye pigment in Drosophila"
                     /translation="MSCPYAGNGNDHDDSAVPLTTEVGKIYGEYLMLDKLLDAQCMLSE
                     EDKRPVHDEHLFIITHQAYELWFKQIIFEFDSIRDMLDAEVIDETKTLEIVKRLNRVVL
                     ILKLLVDQVPILETMTPLDFMDFRKYLAPASGFQSLQFRLIENKLGVLTEQRVRYNQKY
                     SDVFSDEEARNSIRNSEKDPSLLELVQRWLERTPGLEESGFNFWAKFQESVDRFLEAQV
                     QSAMEEPVEKAKNYRLMDIEKRREVYRSIFDPAVHDALVRRGDRRFSHRALQGAIMITF
                     YRDEPRFSQPHQLLTLLMDIDSLITKWRYNHVIMVQRMIGSQQLGTGGSSGYQYLRSTL
                     SDRYKVFLDLFNLSTFLIPREAIPPLDETIRKKLINKSV"
     protein_bind    5640..5709
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"