Very detailed and well summarized. Covered most of the important considerations/options of RNA-seq projects!
The Core welcomed Patricia, an internship student, for the summer 2015!
Just within the 2 weeks of her commitment, she learned various genomics techniques from RNA extraction to HiSeq runs, and now she is working on a single cell RNA-seq project! Tophat is computing overnight and we (hope) will see some cool biology within a couple of days!
All NGS users are encouraged to attend
May 27, 2015, 10:00-4:00
Instructor: Anton Nekrutenko, Penn State University
Hershey, Pennsylvania, United States
Registration is free. Affiliation with Penn State is not required, unless the workshop is oversubscribed, in which case preference will be given to registrants from Penn State. Capacity is limited so you are encouraged to register now.
Are you a biomedical researcher who needs to do complex analysis on large datasets?
Galaxy is an open, web-based platform for data intensive life science research that enables non-bioinformaticians to create, run, tune, and share their own bioinformatic analyses.
This hands-on workshop will teach participants how to integrate data, and perform simple and complex analysis within Galaxy in domains such as variation, RNA-Seq / differential expression, and metagenomics. It will also cover data visualization and visual analytics, and how to share and reuse your bioinformatic analyses, all from within Galaxy.
No programming or Linux command line experience is required.
RNA sequencing reveals a slow to fast muscle fiber type transition after olanzapine infusion in rats.
Second generation antipsychotics (SGAs), like olanzapine, exhibit acute metabolic side effects leading to metabolic inflexibility, hyperglycemia, adiposity and diabetes. Understanding how SGAs affect the skeletal muscle transcriptome could elucidate approaches for mitigating these side effects. Male Sprague-Dawley rats were infused intravenously with vehicle or olanzapine for 24h using a dose leading to a mild hyperglycemia. RNA-Seq was performed on gastrocnemius muscle, followed by alignment of the data with the Rat Genome Assembly 5.0. Olanzapine altered expression of 1347 out of 26407 genes. Genes encoding skeletal muscle fiber-type specific sarcomeric, ion channel, glycolytic, O2- and Ca2+-handling, TCA cycle, vascularization and lipid oxidation proteins and pathways, along with NADH shuttles and LDH isoforms were affected. Bioinformatics analyses indicate that olanzapine decreased the expression of slower and more oxidative fiber type genes (e.g., type 1), while up regulating those for the most glycolytic and least metabolically flexible, fast twitch fiber type, IIb. Protein turnover genes, necessary to bring about transition, were also up regulated. Potential upstream regulators were also identified. Olanzapine appears to be rapidly affecting the muscle transcriptome to bring about a change to a fast-glycolytic fiber type. Such fiber types are more susceptible than slow muscle to atrophy, and such transitions are observed in chronic metabolic diseases. Thus these effects could contribute to the altered body composition and metabolic disease olanzapine causes. A potential interventional strategy is implicated because aerobic exercise, in contrast to resistance exercise, can oppose such slow to fast fiber transitions.
- [PubMed – in process]
Finally I’m updating my blog as well as the GS core web site with lots of new information on our currently available services and prices. Glad I finally came here! We keep growing and we will be able to accommodate any genomics services you might have in your mind. Just give me a shout with your ideas!
Genome Sciences Core Facility
Summary Statement: (last updated 1/16/15)
The Penn State Hershey Genome Sciences Facility is a full service facility and provides consultation, instrumentation, and services to both Penn State and non-Penn State investigators in genomic, epigenomic, and transcriptomic studies (http://www.pennstatehershey.org/web/core/gene-expression-analysis-overview). The variety of instrumentation allows for capabilities ranging from highly focused analysis of candidate SNPs, and mRNAs to whole genome, exome, epigenome, and transcriptome sequencing. Services are also available for a variety of study designs extending from a few laboratory samples to large (100s to 1,000s of samples) clinical projects. The full bioinformatics service is also available for data analysis.
The Facility resides in 5,000 sq ft of newly renovated space, encompassing separate “pre-amplification” and “post-amplification” rooms to prevent any contamination of PCR-amplified materials to pre-processed input DNA/RNA samples. Four well-experienced staffs are available for assisting project operation. In addition, the lab space is available for investigators who need temporary room for sample preparation.
We receive either tissue, DNA/RNA, or customer-generated NGS libraries. We process samples according to the prior consultation and agreement on the design of experiment. We develop new applications to accommodate state-of-the-art NGS technologies. We conduct sequencing reads alignment, secondary analysis (quantitation, variant calling, functional annotation, visualization, etc) and follow-up the interpretation of the results. We support grant writings and educate/train students/post-docs for hands-on NGS processing.
For the most up-to-date information, see the facility’s web page and links at http://med.psu.edu/web/core/gene-expression-analysis.
Facility Details: (last updated 1/16/15)
Nucleic acid processing and analysis: The Facility maintains equipment for nucleic acid extraction (QIAsymphony and Bullet Blender), sonication (Covaris adaptive focused acoustics ultrasonicator E-series), quantitation and quality control (Agilent 2100 Bioanalyzer, Nanodrop Spectrophotometer, Qubit, and access to a Molecular Devices FlexStation 3 UV/Vis/Fluor spectrophotometer). An LifeTech QuantStudio 12K Flex system with robotic plate handler and QuantStudio 3D digital PCR system is available for qPCR gene expression and genotyping services. The Facility provides custom Sanger sequencing of plasmid, PCR and BAC templates. We can also run fragment analysis assays including footprinting, microsatellite, loss of heterozygosity (LOH), SSCP, RFLP and genotyping/SNPlex (multiplexed SNP analysis). Cost for each service is as follows (we allow walk-up and free usage of Nanodrop and Qubit, please inquire for the usage of FlexStation 3) :
|Sample prep and PCR service||for base cost -1 sample||cost per additional sample|
|Bioanalyzer (DNA, RNA, protein) (up to 11-12 samples)||$60|
|Usage of QuantStudio 12K Flex||$14.00|
|Kapa library qPCR||$33.19||$1.52|
|QIAsymphony service||cost per sample|
|Whole Blood DNA prep 200||$5.90|
|Whole Blood DNA prep 400||$7.40|
|Whole Blood DNA prep 1000||$10.80|
|DNA prep buffy coat 200||$5.90|
|DNA prep buffy coat 400||$7.40|
|RNA prep 400||$6.40|
|RNA prep 800||$12.00|
|Saliva DNA prep 1000||$10.80|
|Sanger sequencing||cost per sample|
Microarray: The Facility offers Illumina platforms as the main microarray-based procedure for genotyping and gene expression. The recently acquired iScan system allows high-throughput and cost-effective microarray analysis of genotyping, gene expression, copy number analysis, methylation analysis and LOH studies. Roche and Agilent microarray equipment is also available. Individual SNP calls can be confirmed by PCR tools described above. Listed below are the costs of some popular microarrays.
|Microarray service||minimum sample number required||cost per sample|
|Infinium Omni 2.5 (16 samples)||16||214.13|
|Infinium Omni 2.5 (48 samples)||48||200.63|
|Human Omni 5||16||319.17|
|Human Omni Express-24 v1.0||48||149.65|
|Human gene expression||24||98.72|
|Mouse gene expression||24||132.65|
Next Generation Sequencing platforms: For next generation sequencing projects the Facility offers a range of workflows depending on investigator needs and number of reads/length of sequence required for the project. The Facility contains a MiSeq system to perform low output applications (from 300 Mb to 7.5 Gb in ~4 to 39 hours) and a HiSeq 2500 system, which allows data collection in both rapid run (up to 90 Gb/flow cell, 2×150 bp, in 40 hours) and high output modes (up to 300 Gb/flow cell, 2×100 bp, in ~11 days) on either one or two flow cells. The Facility also maintains a LifeTech Ion Proton DNA sequencer for targeted sequencing and exome sequencing (10 Gb with sequencing run time of 2-4 hours). An Apollo 324 robotic system supports a variety of automated library preparation protocols for whole genome, whole exome, ChIP-seq, RNA-seq and targeted resequencing. Listed below are currently operated services, but we are expanding the service to meet the needs of each investigator.
|NGS library prep service||cost per sample*|
|whole genome sequencing||WGS library prep (minimum input DNA 100 ng)||$50.12|
|whole exome sequencing||exome (Human) (minimum input DNA 1 ng for intact DNA, under R&D for FFPE DNA)||$362.18|
|exome (other species, mouse, bovine, zebrafish)||please inquire|
|ChIP sequencing||ChIP-seq library prep (minimum input DNA 0.5 ng)||$50.12|
|Low input ChIP-seq library prep (minimum input DNA 0.05 ng)||$115.33|
|Methylation sequencing||ERRBS (enhanced reduced representation of bisulfite sequencing) library prep||$99.33|
|RNA sequencing||PolyA RNA seq library prep (strand-specific, minimum input RNA 50 ng)||$69.01|
|Total RNA seq library prep (rRNA depleted, human, mouse, rat, standard rRNA-depletion, strand-specific, minimum input RNA 100 ng)||$160.55|
|Low Input RNA-seq library prep (minimum input RNA 10 pg, or single cell)||$80.88|
|Low Input cDNA synthesis (minimum input RNA 10 pg, or single cell)||$70.35|
|Degraded Low Input RNA-seq library prep (minimum input RNA 10 ng)||$194.58|
|Degraded Low Input cDNA synthesis||$144.46|
|Small RNA seq library prep (minimum input RNA 1 ug)||$96.96|
|Low input small RNA-seq library prep (minimum input RNA 100 ng)||$114.97|
|Single cel RNA-seq (up to 96 samples)||please inquire|
|*QC bioanalyzer run will be added (usually 3 runs ($180) up to 11 samples)|
|Seq only||2X: paired end, 1X: single end|
|Rapid per 2 lanes (need 2 lanes to run the flow cell)||Reagent kit||No. of reads (million) per flow cell (2 lanes run mixed/run together)||cost|
|Rapid per lane (need 2 lanes to run the flow cell, cBot duo to separate lane 1 and 2)||Reagent kit||No. of reads per lane||cost|
|Output per lane (need 8 lanes to run the flow cell)||Reagent kit||No. of reads per lane||cost|
|versions||MiSeq reagent kit||No. of reads (million)||cost|
|2X150 micro (small genome)||4||$1,006.31|
|2X150 nano (small genome, QC)||1||$746.39|
Bioinformatics Support: The Penn State College of Medicine has three dedicated High Performance Computing (HPC) systems with a total of 136 CPU Cores, 768GB RAM and 50TB of local storage in addition to the 80 TB Storage Area Network (SAN). For the non-human subject samples, we also have access to the world class High Performance Compute systems at Penn State University Park, including BioSTAR (512 processor cores, 4 TB of aggregate memory, and 624 TB of disk storage) and CyberSTAR (1,920 cores and 7.68 TB memory, and 650 TB storage). We conduct sequencing reads alignment, secondary analysis (quantitation, variant calling, functional annotation, visualization, etc), as well as statistical analysis correlating genomic with phenotypic information. There is no charge for the consultation and we encourage each investigator to make an appointment with Director/staffs in advance to secure enough time for discussion. Director’s availability is found in the 4th tab of below web site(http://sites.psu.edu/yuka/). Bioinfomatics service will be charged at $60 per hands on hour. Each project is different, so investigators are encouraged to contact us to estimate the necessary hours for each project.
|Bioinformatics support||cost per hour|
For the upcoming holiday weeks, Genome Sciences Facility will be staffed on 12/22, 12/23, 12/24 and 12/29, 12/30, 12/31.
There will be no staff available Thursdays or Fridays either week.
Regular hours will resume the week of January 5.
Genewiz Sanger sequencing service will not be available for pick up on 12/25 and 1/1. Anything shipped on the 26th or the 2nd will not be processed until MONDAY as they do not accept packages on Saturdays.
Some may choose to wait and ship their packages on Monday so they do not sit with FedEx over the weekend.
I myself will be in office except 12/24 morning, 12/25, and 1/1, so I’m happy to assist Genewiz shipping if anyone wishes to ship out on 26th or 2nd.
Thank you and I’m wishing you safe and happy holidays!
I’m happy to find this site finally:
Robust and easy to use!
I am away from Mon (12/8) to Fri (12/12).
If you need immediate assistance, please contact the Core staffs.
GS core will be closed on Thursday, Nov. 27 and Friday, Nov. 28 . Genewiz will receive the samples submitted on Nov. 26 (Wed) on Nob. 28 (Fri) and return the data within the day.
We are working hard to get 2 HiSeq Rapid runs started this evening knowing there will be a first snow storm here tomorrow!
Please stay safe and warm…