Non-coding RNAs (ncRNAs) have emerged as versatile master regulators of biological functions in recent years. Long ncRNAs (lncRNAs) are a large and diverse class of transcribed ncRNAs whose lengths exceed that of 200 nucleotides. There has been a breakthrough in genome editing technology, the CRISPR-Cas9 (clustered regulatory interspaced short palindromic repeats/CRISPR-associated protein 9) technology, in the last decade. CRISPR loci are transcribed into ncRNA and eventually form a functional complex with Cas9 and further guide the complex to cleave complementary invading DNA.

Lifeasible provides functional analysis of plant lncRNA with CRISPR-Cas9 technology, successfully applied in model plants such as Arabidopsis and tobacco and essential crops like wheat, maize, and rice. Our platform is equipped with cutting-edge facilities and professional experts to support research. Here, we provide various services according to customers' demands.

Cloning of the Full-Length lncRNA

We help our customers to perform RNA extraction, real-time RT-PCR, and cloning of the full-length lncRNA. Firstly, we clone the 5' 7-methyl guanylate cap and 3' poly A tail sequences. Then, primers are designed to amplify 5' and 3' sequences based on the RNA-seq sequence of lncRNA. Last, the amplified fragment is finally determined by Sanger sequencing.

CRISPR/Cas9 Vector Construction

Fig. 1 Series of events to generate a clustered regulatory interspaced short palindromic repeats/CRISPR-associated (CRISPR-Cas) mutagenized plant.

• The small size of the guide RNA (20 bp) allows the co-delivery of multiple 'single guide' RNAs (sgRNA) with Cas9 to the cell, making it feasible to simultaneously edit more than one target sequence at the same time. The ease and robustness of this technology make it an attractive genome editing tool for plant biology.
• We provide target selection and plasmid construction services. Specific gRNA targeting lncRNA is selected using the online tool CRISPR-P. To obtain high editing efficiency, the GC% in specific target sequences should be between 40% and 70%. In addition, four or more consecutive T nucleotides in the target sequence should be avoided, and there should be 5 bp between the target sequence and the guide RNA (gRNA) sequence. The amplified fragment is assembled on the binary plasmid through the Golden Gate ligation method.

Plant Genetic Transformation

We help our customers deliver the CRISPR/Cas 9 vector into plants via different systems, such as Gemini virus-mediated delivery, agrobacterium transformation, and protoplast transformation.

Lifeasible provides cost-effective, high-quality, and hassle-free services to our customers worldwide. We provide our clients with direct access to our experts and prompt responses to their questions. If you are interested in our services or have questions, please contact us or make an online inquiry.

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For research or industrial use.

Related Services

• Plant lncRNA Gain-of-Function Analysis
• Plant lncRNA Loss-Of-Function Analysis
• Plant lncRNA Other Knockdown Analysis