PMC:3577827 / 29423-30996 JSONTXT

{"project":"PGDBj_disease_curation1","denotations":[{"id":"T252","span":{"begin":294,"end":301},"obj":"3711"},{"id":"T253","span":{"begin":303,"end":311},"obj":"3708"},{"id":"T254","span":{"begin":316,"end":324},"obj":"3659"},{"id":"T255","span":{"begin":403,"end":409},"obj":"4081"},{"id":"T256","span":{"begin":1208,"end":1215},"obj":"3711"},{"id":"T257","span":{"begin":1228,"end":1236},"obj":"3708"},{"id":"T258","span":{"begin":1266,"end":1274},"obj":"3659"},{"id":"T259","span":{"begin":1289,"end":1295},"obj":"4081"},{"id":"T260","span":{"begin":76,"end":80},"obj":"At5g45250"},{"id":"T261","span":{"begin":85,"end":89},"obj":"At5g45260"},{"id":"T262","span":{"begin":431,"end":435},"obj":"At5g45250"},{"id":"T263","span":{"begin":440,"end":444},"obj":"At5g45260"},{"id":"T264","span":{"begin":47,"end":57},"obj":"http://purl.obolibrary.org/obo/ECO_0000315"},{"id":"T265","span":{"begin":205,"end":208},"obj":"http://purl.obolibrary.org/obo/ECO_0000008"},{"id":"T266","span":{"begin":283,"end":293},"obj":"http://purl.obolibrary.org/obo/ECO_0000315"},{"id":"T267","span":{"begin":343,"end":353},"obj":"http://purl.obolibrary.org/obo/ECO_0000315"},{"id":"T268","span":{"begin":392,"end":402},"obj":"http://purl.obolibrary.org/obo/ECO_0000315"},{"id":"T269","span":{"begin":669,"end":672},"obj":"http://purl.obolibrary.org/obo/ECO_0000008"},{"id":"T270","span":{"begin":703,"end":706},"obj":"http://purl.obolibrary.org/obo/ECO_0000008"},{"id":"T271","span":{"begin":837,"end":840},"obj":"http://purl.obolibrary.org/obo/ECO_0000008"},{"id":"T272","span":{"begin":878,"end":881},"obj":"http://purl.obolibrary.org/obo/ECO_0000008"}],"text":"Expression Analysis of Defense-related Gene in Transgenic Plants Expressing RPS4 and RRS1 \nExpression of the pathogenesis-related 1 (PR1) gene involved in the plant defense responses was determined by qRT-PCR of RNA from five leaf-disks cut from leaves of the 2.5 true leaf stage T2 transgenic B. rapa, B. napus and cucumber, four-week-old T3 transgenic N. benthamiana, and three-week-old T2 transgenic tomato plants carrying both RPS4 and RRS1 using a cork borer (No. 3). Total RNA was isolated and treated with RNase-free DNase (Promega, WI, USA). 500 ng of total RNA was synthesized with oligo dT primer using a PrimeScript RT reagent kit (Takara, Otsu, Japan). QRT-PCR was performed with SYBR Green PCR Master Mix (BIO-Rad Laboratories, CA, USA) using the first-strand cDNA as a template on an MJ Opticon (Bio-Rad Laboratories). QRT-PCR mixtures consisted of 1xSYBR Green I PCR Master Mix and 200 nM (each) sense and antisense primers. Following a preliminary denaturation step at 95°C for 30 s, the reaction mixtures were cycled 40X at 95°C for 5 s and at 65°C for 20 s. The target sample copy number was averaged for two reactions, and the experiment was repeated twice. Expression of the Br-CBP20 for B. rapa, Bn-ACT for B. napus, EF1α for N. benthamiana and cucumber, or Tip41 for tomato was used for normalization. PR1 gene expression is shown as relative values set at a value of 1 in the control plants. Nucleotide sequences of gene-specific primers for each gene are listed in Table S1. Bars indicate SE. This experiment was repeated twice with similar results."}