Genetic dissection of scent metabolic profiles in diploid rose populations

The scent of flowers is a very important trait in ornamental roses in terms of both quantity and quality. In cut roses, scented varieties are a rare exception. Although metabolic profiling has identified more than 500 scent volatiles from rose flowers so far, nothing is known about the inheritance of scent in roses. Therefore, we analysed scent volatiles and molecular markers in diploid segregating populations. We resolved the patterns of inheritance of three volatiles (nerol, neryl acetate and geranyl acetate) into single Mendelian traits, and we mapped these as single or oligogenic traits in the rose genome. Three other volatiles (geraniol, β-citronellol and 2-phenylethanol) displayed quantitative variation in the progeny, and we mapped a total of six QTLs influencing the amounts of these volatiles onto the rose marker map. Because we included known scent related genes and newly generated ESTs for scent volatiles as markers, we were able to link scent related QTLs with putative candidate genes. Our results serve as a starting point for both more detailed analyses of complex scent biosynthetic pathways and the development of markers for marker-assisted breeding of scented rose varieties.     

Selecting optimal oligonucleotide composition for maximal antisense effect following streptolysin O-mediated delivery into human leukaemia cells.

It is widely accepted that most cell types efficiently exclude oligonucleotides in vitro and require specific delivery systems, such as cationic lipids, to enhance uptake and subsequent antisense effects. Oligonucleotides are not readily transfected into leukaemia cell lines using cationic lipid systems and streptolysin O (SLO) is used to effect their delivery. We wished to investigate the optimal oligonucleotide composition for antisense efficacy and specificity following delivery into leukaemia cells using SLO. For this study the well characterised chronic myeloid leukaemia cell line KYO-1 was selected and oligonucleotides (20mers) were targeted to an empirically identified accessible site of c- myc mRNA. The efficiency and specificity of antisense effect was measured 4 and 24 h after SLO-mediated delivery of the oligonucleotides. C5-propyne phosphodiester and phosphorothioate compounds were found to present substantial non-specific effects at 20 microM but were inactive at 0.2 microM. Indeed, no antisense-specific effect was noted at any concentration at either time. All of the other oligonucleotides tested induced some measurable antisense effect, except 7 (chimeric, all-phosphorothioate, 2'-methoxyethoxy termini) which was essentially inactive at 20 microM. The rank efficiency order of the remaining antisense compounds was 4 = 3 >> 9 >> 10 = 8 = 5 = 6 > 11. The efficient antisense effects induced by the chimeric methylphosphonate-phosphodiester compounds were found to be highly specific. Increased phosphorothioate content in the oligonucleotide backbone correlated with reduced antisense activity (efficacy: 2'-methoxyethoxy series 9 >> 8 >> 7, 2'-methoxytriethoxy series 10 > 11). No consistent evidence was obtained for increased activity correlating with increased oligonucleotide-mRNA heteroduplex thermal stability. In conclusion, the chimeric methylphosphonate-phosphodiester oligodeoxynucleotides present the most favourable characteristics of the compounds tested, for efficient and specific antisense suppression of gene expression following SLO-mediated delivery.