An artificial miRNA as a new tool to silence and explore gene functions in apple

AbstractWe focus this report on the nucleus of the lateral olfactory tract (NLOT), a superficial amygdalar nucleus receiving olfactory input. Mixed with its Tbr1-expressing layer 2 pyramidal cell population (NLOT2), there are Sim1-expressing cells whose embryonic origin and mode of arrival remain unclear. We examined this population with Sim1-ISH and a Sim1-tauLacZ mouse line. An alar hypothalamic origin is apparent at the paraventricular area, which expresses Sim1 precociously. This progenitor area shows at E10.5 a Sim1-expressing dorsal prolongation that crosses the telencephalic stalk and follows the terminal sulcus, reaching the caudomedial end of the pallial amygdala. We conceive this Sim1-expressing hypothalamo-amygdalar corridor (HyA) as an evaginated part of the hypothalamic paraventricular area, which participates in the production of Sim1-expressing cells. From E13.5 onwards, Sim1-expressing cells migrated via the HyA penetrate the posterior pallial amygdalar radial unit and associate therein to the incipient Tbr1-expressing migration stream which swings medially past the amygdalar anterior basolateral nucleus (E15.5), crosses the pallio-subpallial boundary (E16.5), and forms the NLOT2 within the anterior amygdala by E17.5. We conclude that the Tbr1-expressing NLOT2 cells arise strictly within the posterior pallial amygdalar unit, involving a variety of required gene functions we discuss. Our results are consistent with the experimental data on NLOT2 origin reported by Remedios et al. (Nat Neurosci 10:1141–1150, 2007), but we disagree on their implication in this process of the dorsal pallium, observed to be distant from the amygdala.

Download Full-text

Optimisation of tobacco rattle virus-induced gene silencing in Arabidopsis

Functional Plant Biology ◽

10.1071/fp05096 ◽

2006 ◽

Vol 33 (4) ◽

pp. 347 ◽

Cited By ~ 46

Author(s):

Changchun Wang ◽

Xinzhong Cai ◽

Xuemin Wang ◽

Zhong Zheng

Keyword(s):

Gene Silencing ◽

High Throughput ◽

Function Analysis ◽

Tobacco Rattle Virus ◽

Phytoene Desaturase ◽

Growth Stages ◽

Virus Induced Gene Silencing ◽

Gene Functions ◽

Genes Encoding ◽

Gene Function Analysis

Arabidopsis thaliana (L.) Heynh. is a model plant species in which to study plant gene functions. Recently developed virus-induced gene silencing (VIGS) offers a rapid and high-throughput technique platform for gene function analysis. In this paper we report optimisation of tobacco rattle virus (TRV)-induced gene silencing in Arabidopsis. The parameters potentially affecting the efficiency of VIGS in Arabidopsis were investigated. These included the concentration and pre-incubation of Agrobacterium inocula (agro-inocula), the concentration of acetosyringone included in agro-inocula, the Agrobacterium inoculation (agro-inoculation) method, the ecotypes and the growth stages of Arabidopsis plants for agro-inoculation, and the growth temperature of agro-inoculated plants. The optimised VIGS procedure involves preparing the agro-inocula with OD600 of 2.0, pre-incubating for 2 h in infiltration buffer containing 200 μm acetosyringone, agro-inoculating by vacuum infiltration, and growth of agro-inoculated plants at 22 −24°C. Following this procedure consistent and highly efficient VIGS was achieved for the genes encoding phytoene desaturase (PDS) and actin in Arabidopsis. The silencing phenotype lasts for at least 6 weeks, and is applicable in at least seven ecotypes, including Col-0, Cvi-0, Sd, Nd-1, Ws-0, Bay-0 and Ler. TRV-induced VIGS was expressed not only in leaves, but also in stems, inflorescences and siliques. However, VIGS was not transmissible through seed to the subsequent generation. The optimised procedure of the TRV-induced gene silencing should facilitate high-throughput functional analysis of genes in Arabidopsis.

Download Full-text