Reverse Genetics for Functional Genomics of Phytopathogenic Fungi and Oomycetes
Figure 3
RNAi strategies: (a) conventional hpRNAi, transcripts with complementary or near-complementary 20- to 50-base pair inverted repeats form double-stranded RNA (dsRNA) hairpins. These dsRNAs are processed into micro-RNAs that mediate mRNA degradation. (b) pSilent1 (heterogeneous nuclear RNA expressing vector system), it carries a hygromycin resistance cassette and a transcriptional unit for hairpin RNA expression with multiple cloning sites and a spacer of an intron sequence. (c) pSilent-Dual1 system (opposing dual promoter system), trpC and gpdA promoters were cloned in a convergent manner separated by multicloning site. For co-silencing, a 0.41 kb eGFP fragment was first inserted in pSD1, resulting in pSD1-G. Gene fragments of can be inserted into pSD1 vector, which will express corresponding chimeric double-stranded RNA, a template for homology-dependent degradation of the target mRNA.
(a) Conventional hpRNASystem
(b) pSilent 1(pS1) system (Nakayashiki et al., 2005)
(c) pSilent-Dual1(pSD1)System (Nguyen et al., 2008)
