RNA Sequencing Services
www.lcsciences.com 1-888-528-8818
microRNA / Small RNA Sequencing Service
Next-gen sequencing is a new method and powerful tool
used to identify and quantitatively decode the entire
population of small RNAs in your total RNA sample. LC
Sciences provides a comprehensive sequencing service
utilizing Illumina’s high-throughput sequencing
technology which enables comprehensive, highly
sensitive and specific discovery and profiling of all forms
of small RNAs in your sample.
Single-Cell RNA-Seq Service
Single-cell RNA-Seq can be used to examine the
expression of individual cells and provides a higher
resolution of cellular differences as compared to
traditional RNA-Seq. Single-cell RNA-Seq enables us to
understand the function of an individual cell in the
context of its microenvironment. Start with as little as
10 pg-10 ng of total RNA.
Digital Gene Expression (mRNA) Sequencing Service
The DGE service offered by LC Sciences makes use of
RNA Sequencing technology to “count” each individual
transcript, offering distinct advantages over semi-
quantitative array-based gene-expression analysis
systems which rely on signal intensities. As an
alternative to expression arrays, DGE provides a cost
effective basic differential expression analysis of known
genes via the latest next-gen sequencing technology.
Poly(A) RNA-Seq / Total RNA-Seq Sequencing Services
In addition to expression quantification, RNA-Seq
technology can be used to determine the structure of
genes, their splicing patterns or other post
transcriptional modifications, and to detect rare and
novel transcripts. Total RNA-Seq enables the sequencing
of both poly(A) and non-poly(A) RNAs, such as long non
-coding RNA (lncRNA), via a ribosomal depletion library
preparation strategy. LC Sciences provides a turnkey
solution for transcriptome sequencing featuring the
latest in RNA-Seq technology.
LC Sciences Comprehensive Services
60-03F-02
Key Advantages of RNA Sequencing RNA Sequencing provides a more comprehensive view of the transcriptome. RNA Sequencing enables us to sequence and profile all
species of transcripts in your total RNA samples - from mRNA to non-coding RNAs to small RNAs.
RNA Sequencing is not necessarily dependent on any prior sequence knowledge. There is no need for the design of probes that
must be based on prior sequence or secondary structure information. Therefore, transcriptome profiling in any species is possible, making this
method particularly attractive for non-model species. Additionally, RNA Sequencing data can be used to build de novo gene models.
RNA Sequencing has increased dynamic range and sensitivity. RNA Sequencing enables you to achieve “digital” transcript expression
analysis meaning that expression level data are based on each individual transcript that is sequenced and counted. By increasing the
sequencing depth, a potentially unlimited dynamic range can be reached making RNA Sequencing an ideal tool for detection rare transcripts.
RNA Sequencing provides information of sequence variation in transcripts. RNA-Seq yields a rich data set that includes information
about post transcriptional mutations and their genomic context. Because RNA-Seq data yields information about how exons are connected, it
can reveal sequence variations in the transcripts due to alternative splicing events and provide allele-specific or isoform-specific gene
expression information. Additionally, RNA-Seq data is useful for gene mapping functions such as describing the length of UTRs and exon
boundaries.
Turnkey Solution These comprehensive services are designed to be one-stop and
produce the results needed to quickly advance your biological and
biomedical research. For all types of RNA sequencing, our “Total RNA
to Data” services include: sample prep/QC, library preparation, RNA
sequencing, bioinformatics analysis, and high-level customer support.
Expert Services LC Sciences has been providing RNA discovery, profiling and related
bioinformatics services since 2005 and our experts have examined
thousands of varied RNA datasets, giving us unique insight into RNA
research. We don’t merely deliver data; we will handle all the advanced
bioinformatics analysis and help you find answers to the questions
you’re asking.
Deep Sequencing
Advanced Data Analysis
Library Preparation
microRNA Sequencing Service
60-03F-03
Group 1
Known Pig
miRNAs
Others
Group 2
Known miRNAs
Candidate pig miRNAs
Group 3
Candidate pig miRNAs
genome inconsistent
with miRBase
Group 4
Potentially
novel miRNAs
No Hit
RNA Sequencing provides an excellent tool for discovery of novel microRNAs (miRNAs) as well as genome-wide profiling of small RNA and
miRNA expression in various samples. LC Sciences provides a discovery sequencing service utilizing the Illumina high-throughput sequencing
technology which enables comprehensive coverage, highly sensitive and specific discovery and profiling of all forms of small RNAs in your
sample.
Profile miRNA in any species - there is no need for probes based on prior sequence or secondary structure information.
Profiling is transcriptome-wide - investigate all miRNAs and small RNAs, of any size, known and unknown, in your sample.
Discover novel small RNAs and detect rare transcripts and transcript variants, such as sequence isoforms and SNPs.
microRNA Sequencing Data Flow
Example - pig sample generated ~20M
reads. Sequencing reads are mapped to
pig mirs in miRBase, mirs of related
species and mapped to pig genome.
(713) 664-7087 1-888-528-8818 Fax (713) 664-8181 [email protected] www.lcsciences.com
Bioinformatics Package Includes: Raw sequencing data – up to 100M 50bp single-end reads per
lane – either FASTA-A or FASTA-Q files with base-calling quality
scores
LC Sciences analysis and quality filtering - processed data is
reduced to mappable reads
Custom construction of reference database(s) - miRBase,
genome, etc and mapping of all quality reads
Alignment, classification, & functional annotation of all mapped
reads
Biostatistical analysis – expression analysis, multi-parameter data
analysis, length distribution, transcript copy number
comparisons, etc
miRNA target prediction
miRNA target GO and KEGG enrichment analysis
Digital Gene Expression (mRNA) Sequencing Service
60-03F-05
Test_id Gene Locus FPKM
Sample1
FPKM
Sample2
Log
(fold
change)
Test_stat P_value Significant
XLOC_041924 BCLAF1 1:29312111-29312566 7.23 0.30 -3.18 -3.12 0.0018 Yes
XLOC_145478 MOSPD1 X:108883967-108884284 13.84 4.76 -1.07 -3.10 .0019 Yes
XLOC_150271 IRF7 2:294234-294407 11.27 28.54 .93 3.09 .0020 Yes
XLOC_180169 SORBS2 15:44062921-44063290 8.59 0.37 -3.14 -3.08 .0021 Yes
The incredible power of RNA-Seq was initially applied primarily for complex whole transcriptome applications such as discovery of rare genes,
splice junctions and gene fusions or for the study of novel and less well-characterized organisms. However, as the technology is improving and
costs are decreasing, RNA sequencing is now suitable to be used for simple expression profiling of known genes as well.
An Alternative to Expression Arrays Achieve digital transcript expression analysis – RNA sequencing technology digitally “counts” each individual transcript, offering distinct
advantages over semi-quantitative array-based gene-expression analysis systems which rely on relative signal intensities.
Expression profiling is truly genome-wide – there is no need for probes based on prior sequence information.
Accurate quantitation of expression levels over 5 orders of magnitude of transcript abundance.
Profile gene expression in any species – quantitate differential expression across groups of different biological samples.
Provides a cost effective basic differential expression analysis of known genes via the latest next-gen sequencing technology.
Bioinformatics Package Includes: Raw sequencing data – up to 100M 50bp single-end reads per lane – either FASTA-
A or FASTA-Q files with base-calling quality scores
Data reduced to mappable reads, a list of unique sequences and their copy numbers
Genomic mapping of quality sequencing reads
Gene expression profiling
Test for differential expression at gene and transcript level
GO and KEGG annotation and enrichment analysis
Simple report of analysis showing statistical significance of differential expression
Digital Gene Expression (mRNA) Sequencing Service
60-03F-06
LC Sciences provides a turnkey solution for transcriptome sequencing featuring the latest in RNA-Seq technology. Next-gen sequencing is a
powerful tool used for transcriptome analysis that can determine the structure of genes, their splicing patterns and other post transcriptional
modifications, detect rare and novel transcripts, and quantify their changing expression levels during development and under different disease
conditions.
Profile expression in any species - there is no need for probes based on prior sequence information.
Compare genome-wide expression in different species or otherwise different biological samples.
Detect mutation events such as: novel splicing events, gene fusions, isoforms, SNPs, or other specific coding mutations.
Detect rare, or novel transcripts.
Bioinformatics Package Includes: Raw sequencing data – up to 125M x 2 100bp paired-end reads – either
FASTA-A or FASTA-Q files with base-calling quality scores
Processed data is reduced to mappable reads
De Novo assembly (if applicable)
Alignment of RNA-Seq reads to customer specified reference genome
Identification and construction of splice-junctions
Report of known transcripts with annotation and abundance
Report of novel transcript with abundance
Identification of alternate splicing and report of isoform abundance
Test for differential expression at gene level and transcript level
GO and KEGG annotation and enrichment analysis
Customer data report - includes a summary of methods and all statistical
analysis
RNA-Seq Reads
Align reads to
genome
Assemble transcripts
from spliced alignments
More abundant
Less abundant
Poly(A) RNA-Seq Sequencing Service
(713) 664-7087 1-888-528-8818 Fax (713) 664-8181 [email protected] www.lcsciences.com
Total RNA-Seq (Whole Transcriptome) Sequencing Service
Single-Cell RNA-Seq
60-03F-07
Total RNA-Seq (Whole Transcriptome) sequencing captures a broader range of gene expression changes and enables the detection of novel
transcripts in both coding and non-coding RNA species. Ribo-Zero ribosomal RNA reduction chemistry is used in place a poly-A tail selection
which efficiently removes ribosomal RNA (rRNA). This process minimizes ribosomal contamination and optimizes the percentage of reads
covering RNA species of interest.
Profile expression of any RNA transcript - poly(A) RNA and non-poly(A) RNA.
Obtain annotation and expression information on long non-coding RNA within your samples.
Sequence non-poly(A) prokaryote transcriptomes.
Sequence transcriptomes from degraded RNA from sample types such as formalin-fixed paraffin embedded (FFPE).
Bioinformatics Package Includes: All the analysis provided by the Poly(A) RNA-Seq service, in addition to:
Non-coding RNA information – map to genome, alignment, expression analysis and limited annotation
Optional circular RNA (circRNA) analysis – although the biological function of most circular RNAs is still unclear, some have shown
potential as gene regulators.
Single-cell RNA-Seq can be used to examine the expression of individual cells and provides a higher resolution of cellular differences as
compared to traditional RNA-Seq. Single-cell RNA-Seq enables us to understand the function of an individual cell in the context of its
microenvironment.
Comprehensive Service: Start with as little as 10 pg-10 ng of total RNA
Library preparation – poly(A) tail selection method
Illumina Hi-Seq 2000 – 2 x 100bp paired-end sequencing run
Advanced bioinformatics package
Bioinformatics Package Includes: Illumina base-calling and analysis
Raw sequencing data – 50bp single-end reads – either FASTA-A or FASTA-Q files with base-calling quality scores
LC Sciences quality filtered data reduced to mappable reads, a list of unique sequences and their copy numbers
Genomic mapping of quality sequencing reads
Test for differential expression at gene level.
GO and KEGG annotation and enrichment analysis.
Customer data report – includes a summary of methods and all statistic analysis
Single-Cell RNA-Seq
Degradome Sequencing Service
60-03F-04
Degradome sequencing (Degradome-Seq), also referred to as parallel analysis of RNA ends (PARE), incorporates a modified 5′-rapid
amplification of cDNA ends (RACE) method with high-throughput next-gen sequencing to provide a comprehensive means of analyzing
patterns of RNA degradation.
Because miRNAs can cause endonucleolytic cleavage of mRNA by extensive and often perfect complementarity to mRNAs, degradome
sequencing has been used to identify miRNA cleavage sites. Degradome sequencing has confirmed many known and revealed novel plant
miRNA targets.
In plants, miRNAs tend to cause cleavage of their targets at the position between nucleotides 10 and 11 of the miRNA.
Next-Gen sequencing of the 5’ ends of degraded RNA products allows identification of over-represented 5’ ends (miRNA cleavage sites).
Matching cleavage sites to known miRNA sequences links miRNAs to their targets.
Bioinformatics Package Includes: Raw sequencing data – up to 100M 50bp single-end reads per lane – either FASTA-A or FASTA-Q files with base-calling quality scores
LC Sciences quality filtered data reduced to mappable reads, alignment, classification, & functional annotation of all mapped reads
Identification of rRNAs, tRNAs, snoRNAs, polyN and other non-coding RNAs
Distribution of degradation fragments on selected region of genome
Identification of target mRNAs
Statistical summary of mRNA degradation sites
Identification of miRNAs from miRBase related to mRNA degradation (results will be shown in a target plot diagram)
Customer data report – includes a summary of methods and statistical analysis
Confirmed microRNA targets are presented in the form of target plots (t-plots).
miRNA corresponding to the
target cleavage site
Score, Type, P-value
Reads
mRNA
(713) 664-7087 1-888-528-8818 Fax (713) 664-8181 [email protected] www.lcsciences.com
www.lcsciences.com 1-888-528-8818
RNA Sequencing Services
1. Zhang G et al. (2016) Integrated analysis of mRNA-seq and miRNA-seq in the liver of Pelteobagrus vachelli in response to hypoxia. Sci Rep 6
(1), 22907 [abstract].
2. Han X et al. (2016) Integration of small RNAs, degradome and transcriptome sequencing in hyperaccumulator Sedum alfredii uncovers a
complex regulatory network and provides insights into cadmium phytoremediation. Plant Biotech J 14(6), 1470-1483 [abstract].
3. Chen J et al. (2016) Identification of miRNAs and their targets through high-throughput sequencing and degradome analysis in male and
female Asparagus officinalis. BMC Plant Biol 16(1), 1 [abstract].
4. Ponce D et al. (2016) Tentacle Transcriptome and Venom Proteome of the Pacific Sea Nettle, Chrysaora fuscescens (Cnidaria: Scyphozoa).
Toxins 8(4), 102 [abstract].
5. Sun S et al. (2016) Identification and comparative analysis of the oriental river prawn (Macrobrachium nipponense) microRNA expression
profile during hypoxia using a deep sequencing approach. Comparative Biochemistry and Physiology Part D: Genomics and Proteomics 17, 41-47
[abstract].
6. Wang C et al. (2016) A Group of Novel Serum Diagnostic Biomarkers for Multidrug-Resistant Tuberculosis by iTRAQ-2D LC-MS/MS and
Solexa sequencing. Intl J of Biol Sci 12(2), 246-256 [abstract].
Recent Customer Publications
Description Small/microRNA-Seq
Degradome-Seq Digital Gene Expression
Poly(A) RNA-Seq
Total RNA-Seq Single-Cell RNA-Seq
RNA Selection method
Size Selection – Small RNA
PolyA-tail Selection – mRNA fragments
PolyA-tail Selection – mRNA
PolyA-tail Selection – mRNA
rRNA Depletion – mRNA + lncRNA
PolyA-tail Selection – mRNA
Data Analysis Discovery, Expression, Annotation
miRNA Target ID, Degradation Plots
Expression, Annotation
Discovery, Expression, Annotation
Discovery, Expression, Annotation
Expression, Annotation
Sequencing Instrument
HiSeq HiSeq HiSeq HiSeq HiSeq HiSeq
Max# Samples per Lane
10 10 10 5 5 5
Recommended data size (min reads/sample)
7 million 7 million 7 million 40 million 40 million 20 million
Type of Sample* total RNA total RNA total RNA total RNA total RNA total RNA
Preferred Sample Amt
1 µg 10 µg 3 µg 4 µg 4 µg 10ng
Min Sample Amt Required
10 ng 5 µg 1 µg 2 µg 2 µg 10 pg
Single vs Paired-end Seq Run
Single-end Single-end Single-end Paired-end Paired-end Paired-end
Sequencing Run Length (nts)
50 50 50 2 x 100 2 x 100 2 x 100
@LCSciencesLLC @LCSciences LC Sciences LLC
Inquire about immuno-precipitated RNAs or other enriched RNAs.