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Long-strand non-coding RNA sequencing
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Long-strand non-coding RNA sequencing

Product Description
Case Interpretation

lncRNAs are greater than 200bp in length and do not encode proteins, which are widely found in living organisms. lncRNAs regulate gene expression levels at multiple levels (epigenetic regulation, transcriptional regulation, and post-transcriptional regulation, etc.) and play an important role in life activities. lncRNA sequencing is a research method that uses a specific method to reduce the abundance of rRNAs in samples, during which a library to sequence the enriched lncRNA sequencing would be constructed and almost all the lncRNA information can be obtained at one time,  providing a comprehensive and in-depth analysis of lncRNA classes and functions to get lncRNA related information of specific biological processes (e.g. development, diseases, etc.) quickly and accurately.

 

Technical route:

1

 

technical parameter:

 

Sample type

Sequencing strategy

Lead time

Sample type: Total RNA

Sample concentration: ≥100 ng/μl

Total sample mass: ≥5 μg

Sample purity:OD260/280=1.8~2.2, OD260/230 = 1.8~2.2

Integrity:RIN≥7

Sequencing mode: Illumina/MGI PE150

Sequencing depth: 10G /12G clean data

45 days

 

Product advantages.

a. Cost-effective: one library can obtain both mRNA and lncRNA information;

b.Total solution: one-stop upstream sequencing & downstream functional validation and consulting services;

c. Comprehensive services: standard analysis, advanced analysis and multi-omics association analysis are all available.

 

Case 1 Characterization and identification of lncRNA in maize

Background

lncRNAs play an important regulatory role in gene expression at multiple levels in eukaryotes, but the amount, characteristics and genetic pattern of expression of lncRNAs in maize remain largely unknown at present

Study

By analyzing maize EST, whole genome, and RNA-seq data, 20,163 lncRNAs were predicted from them, and it was hypothesized that more than 90% of them might be precursors of small RNAs, and 1704 were highly plausible lncRNAs. RNA-Seq data results from 13 different tissues and 105 maize recombinant self-incompatibility lines showed that lncRNAs have a strong of tissue specificity. This study provides a unique resource of maize genome annotation and genome-wide characterization of maize lncRNAs, and the genetic pattern of maize lncRNA expression pattern was obtained using eQTL analysis.

Results

The highly plausible 1704 lncRNAs were found to have an average length of 463 bp, and the lncRNAs contained fewer exons compared to functional genes, with 81% containing only one exon.

RNA-seq results in 13 tissues and 105 RILs materials showed that more than 50% of these lncRNAs had tissue expression specificity.

The eQTL localization analysis revealed that lncRNAs in maize are more regulated by trans than cis.

 

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图1. lncRNA在玉米的10条染色体的分布

 

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Figure 2. Location of lncRNA eQTL in maize seeds

 

Case 2 PVT1 promotes gallbladder cancer proliferation by inhibiting miR-18b-5p through DNA methylation

Background

Gallbladder cancer (GBC) is the most common biliary tract cancer, and the lack of early diagnosis, abnormal invasiveness, and chemoresistance has led to its poor overall prognosis with a 5-10% 5-year survival rate. With the rapid development of sequencing technology, a novel type of regulatory genes with lengths exceeding 200 nucleotides, long noncoding RNAs (lncRNAs), have been recognized to play important biological roles in physiological and pathological development. In recent years, there is increasing evidence that dysregulation of lncRNAs is involved in cancer progression.

Study

The novel lncRNA PVT1 plays an important role in the progression of GBC as an oncogene. PVT1 is upregulated in GBC tissues and cells, and its upregulation is associated with poor prognosis in GBC patients. PVT1 promotes GBC cell proliferation and increases tumorigenicity in nude mice. In addition, miR-18b-5p down-regulated by PVT1 was screened by miRNA microarray analysis. Mechanistically, PVT1 promotes miR-18b-5p DNA promoter methylation by recruiting DNMT1 through EZH2 and then represses its expression.

Results.

lncRNA PVT1 expression was upregulated in  GBC tissues and cells, and high PVT1 expression correlated with the prognosis of GBC patients. lncRNA PVT1 knockdown inhibited GBC cell proliferation in vitro and in vivo. In addition, PVT1 facilitated DNA promoter methylation of miR-18b-5p through recruitment of DNMT1 by EZH2, which in turn attenuated the inhibitory effect of miR-18b-5p on HIF1A and ultimately mediated GBC progression.

 

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Figure 1: miR-18b-5p down-regulates HIF1A and promotes GBC cell proliferation in vitro

 

 reference

  1. Li L, Eichten S R, Shimizu R, et al. Genome-wide discovery and characterization of maize long non-coding RNAs[J]. Genome Biology, 2014, 15(2):R40.

  2.Jin, L., Cai, Q., Wang, S. et al. Long noncoding RNA PVT1 promoted gallbladder cancer proliferation by epigenetically suppressing miR-18b-5p via DNA methylation[J]. Cell Death Dis 11, 871 (2020).

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