miRNAs play an important role in plant development, disease resistance, and stress response. It is believed that these small non-coding RNAs modulate plant gene expression at the post-transcriptional level by targeting mRNA cleavage. Targeted changes in the expression of these genes are regulated by differential expression of hundreds of mature miRNAs, each of which binds to a specific sequence of the target mRNA. Therefore, analysis of plant miRNA regulation and expression can provide valuable information for plant tissue development, drought protection and disease defense mechanisms. At present, it is basically impossible to analyze mature miRNAs of plants by qRT-PCR. Due to the characteristics of plant miRNAs, the reaction efficiency based on polyadenylation reverse transcription reaction is very low. QIAGEN has launched a unique solution for universal reverse transcription of mature miRNAs in plant tissues.

QIAGEN miScript Plant RT Kit is used for cDNA synthesis of plant miRNAs and other 3'-modified small RNAs; it is universal for unbiased cDNA synthesis of small RNAs with 3'-end methylation (2'-O-Me) modification Type kit. Designed and optimized for small RNA expression analysis applications based on real-time quantitative PCR, this kit can be used with miScript SYBR Green PCR Kits and miScript miRNA PCR Arrays or miScript Primer Assays. This class of functionally validated analytical and chip products can be applied to the detection of rice, corn, soybeans, tobacco and other important agricultural crops, as well as botany-related research. Small RNAs with 2'-O-Me methylation modification at the 3' end of the base (such as plant mature miRNA and piwi interaction RNA (piRNA)) generally do not have a poly A tail structure and therefore cannot pass through polygland The glycoside-dependent cDNA synthesis method is effective for reverse transcription. The dedicated miScript Plant RT Kit solves this problem by first attaching a linker to the 3' end of the plant miRNA. Unlike polyadenylation, this linkage reaction is not negatively inhibited by the 2'-O-Me methylation modification present at the 3' end of the plant miRNA. Once the linker is successfully connected, a reverse transcription reaction can be achieved.

The combination of the miScript Plant RT Kit and the miScript Plant qPCR System ensures sensitive and specific miRNA detection and quantification for plant samples. Because of the low abundance of many miRNAs, detection sensitivity is critical to ensure reliable data. The miScript Plant qPCR System delivers excellent sensitivity, a wide dynamic range of monitoring, and consistently high amplification efficiency. The good performance of the miScript Plant qPCR System will help you achieve highly repeatable results between the first iterations of technology and bio-repetitive samples.

The reverse transcription reaction can be conveniently carried out. Generally, in the ligation-mediated reverse transcription reaction, total RNA containing a 3'-end 2'-O-Me modified small RNA is generally used as a starting material. Reverse transcription is a straightforward procedure involving incubation for 1 hour at 37 °C and subsequent inactivation of the reaction by brief incubation at 95 °C.

The miScript Plant RT Kit is typically used as part of the miScript Plant PCR System for:

• Quantitative and expression profiling of plant mature miRNAs
• Quantitative and expression profiling of piRNA

The above information source QIAGEN:

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