The following information accompanied stocks donated to the Bloomington Stock Center by Omar Akbari, University of California, San Diego.
PBac{nos-Cas9.T2A.GFP} expresses a 'Cas9.T2A.GFP' cassette, which consists of human codon-optimized Streptococcus pyogenes Cas9 (with a nuclear localization signal at each end) separated by a self-cleaving T2A motif from the GFP open reading frame. Expression is driven by nos promoter sequences. The construct also contains a DsRed marker, expressed from the ubiquitous OpIE2 promoter (derived from the baculovirus Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus).
PBac{nos-Cas9.T2A.GFP}VK00033 is an insertion into the PBac{y+-attP-3B}VK00033 target site at 65B.
PBac{Ubi-Cas9.T2A.GFP} expresses the 'Cas9.T2A.GFP' cassette described for PBac{nos-Cas9.T2A.GFP}, except that expression is driven by a Ubiquitin promoter. The OpIE2-DsRed marker described for PBac{nos-Cas9.T2A.GFP} is also present.
PBac{Ubi-Cas9.T2A.GFP}VK00033 is an insertion into the PBac{y+-attP-3B}VK00033 target site at 65B.
PBac{vas-Cas9.T2A.GFP} expresses the 'Cas9.T2A.GFP' cassette described for PBac{nos-Cas9.T2A.GFP}, except that expression is driven by vas promoter sequences. The OpIE2-DsRed marker described for PBac{nos-Cas9.T2A.GFP} is also present.
PBac{vas-Cas9.T2A.GFP}VK00033 is an insertion into the PBac{y+-attP-3B}VK00033 target site at 65B.
P{U6-Sxl.gRNA} expresses a gRNA that targets Sxl, under the control of the U6-3 promoter.
P{U6-Sxl.gRNA}attP1 is an insertion into the P{CaryP}attP1 target site at 55C.
P{U6-tra.gRNA} expresses a gRNA that targets tra, under the control of the U6-3 promoter.
P{U6-tra.gRNA}attP1 is an insertion into the P{CaryP}attP1 target site at 55C.
P{U6-dsx.F.gRNA} expresses a gRNA that targets the dsx female-specific isoform, under the control of the U6-3 promoter.
P{U6-dsx.F.gRNA}attP1 is an insertion into the P{CaryP}attP1 target site at 55C.
P{U6-betaTub85D.gRNA} expresses a gRNA that targets betaTub85D, under the control of the U6-3 promoter.
P{U6-betaTub85D.gRNA}attP1 is an insertion into the P{CaryP}attP1 target site at 55C.
P{U6-Sxl.gRNA,U6-betaTub85D.gRNA} expresses two gRNAs, targeting Sxl and betaTub85D respectively. Each is expressed under the control of a U6-3 promoter.
P{U6-Sxl.gRNA,U6-betaTub85D.gRNA}attP1 is an insertion into the P{CaryP}attP1 target site at 55C.
P{U6-tra.gRNA,U6-betaTub85D.gRNA} expresses two gRNAs, targeting tra and betaTub85D respectively. Each is expressed under the control of a U6-3 promoter.
P{U6-tra.gRNA,U6-betaTub85D.gRNA}attP1 is an insertion into the P{CaryP}attP1 target site at 55C.
P{U6-dsx.F.gRNA,U6-betaTub85D.gRNA} expresses two gRNAs, targeting the dsx female-specific isoform and betaTub85D respectively. Each is expressed under the control of a U6-3 promoter.
P{U6-dsx.F.gRNA,U6-betaTub85D.gRNA}attP1 is an insertion into the P{CaryP}attP1 target site at 55C.
M{betaTub85D-GFP} expresses GFP under the control of the betaTub85D promoter.
M{betaTub85D-GFP}ZH-86Fa is an insertion into the M{3xP3-RFP.attP'}ZH-86Fa target site at 86E.
P{OpIE2-neoR.traF,OpIE2-pacR.dsxM} encodes two different antibiotic resistance genes, each expressed under the control of the the OpIE2 promoter, with the addition of sex-specific intron and splice site sequence to each coding cassette so that the construct can be used for sex-selection. The 'OpIE2-neoR' fragment encodes neomycin phosphotransferase (neoR), with the addition of the female-specific intron and splice sites from tra so that neoR is only expressed in females. The 'OpIE2-pacR' segment encodes puromycin N-acetyltransferase (pacR), with the addition of the male-specific intron and splice sites from dsx so that pac is expressed only in males. The construct also contains a DsRed marker, expressed under the control of a baculovirus Hr5 enhancer and IE1 promoter.
P{OpIE2-neoR.traF,OpIE2-pacR.dsxM}3 is a homozygous viable and fertile insertion on the third chromosome.