affy_ath_2012_05-Arabidopsis Ubiquitin-Like Proteases characterization

affy_ath_2012_05 - affy_ath_2012_05 - The main objective is to determinate the influence of Ubiquitin-Like Proteases in plant transcriptional modulation. The experiment will also allow us to find what transcriptional cues are particularly affected in the mutants relatively to wild-type.-Seeds were stratified for 3 days at 4ºC in the dark. Afterwards, seeds were surface sterilized, sown onto MS media and grown at 23ºC, with 100 µE light condition, in long days (16h L/8h D). Each MS plate was dived in four, corresponding to a genotype (Col, AB, SIZ1 or SAB), and 10 seeds were sown per genotype. After 10 days, seedlings were collected and frozen in liquid nitrogen. Each replicate was made of ~40 seedlings coming from 4 different MS plates. Frozen plant material was grounded in an eppendorf tube using a mini pestle. RNA was extracted using RNeasy Plant Mini kit (QIAGEN cat.74904), following the manufacturer instructions (QIAGEN, Hilden, Germany). The RNA samples were DNase treated (Recombinant DNase I, Takara Biotechnology, Dalian, China) and subsequently cleaned by column using RNeasy Plant Mini kit (QIAGEN, Hilden, Germany). 12 arrays - ATH1; gene knock out Col [wild type] AB [knock out / insertional mutagen /T-Dna] SIZ1 [knock out / insertional mutagen /T-Dna] SAB [knock out / insertional mutagen /T-Dna]