Guidelines for Submitting Samples to the GSF

Samples for TruSeq Library Preparation

RNA

The GSF requires that total RNA be submitted for mRNA and small RNA sequence analysis. It is recommended that total RNA be purified using the TRIzol® Reagent with Phase Lock Gel™ Heavy procedure (Trizol/Phase Lock Gel Protocol). If retention of miRNA is not required, the Qiagen RNeasy or other comparable isolation kit is acceptable. DNase treatment of RNA samples is strongly recommended to ensure accurate RNA quantitation. Submit RNA samples in nuclease-free H2O at the following concentrations:

  • mRNA-seq: 100 - 250 ng/ul,  1-2ug Total RNA.                      Note: Extra sample is appreciated for all RNA-seq library preparation procedures
  • Stranded mRNA-seq: 100 - 250 ng/ul, 1-2ug Total RNA
  • Stranded Total RNA-seq: 100 - 250 ng/ul, 1-2ug Total RNA
  • small RNA-seq: 250 - 500 ng/ul, 2-3ug Total RNA.

The GSF requires that all RNA samples submitted for sequence analysis be assayed spectrophotometrically. All RNA samples submitted will be subjected to an Agilent 2100 Bioanalyzer quality control run. Submit samples in 1.5 -1.7ml microcentrifuge tubes with the name clearly labeled on the top of the tube with permanent marker as it is written on the order form. Please limit sample names to eight characters.

gDNA / ChIP DNA

The GSF recommends that genomic DNA (gDNA) submitted for sequence analysis be purified using the QIAamp DNA kit or comparable gDNA isolation kits. RNase treatment of gDNA is required. Submit gDNA samples in nuclease-free H2O or reduced EDTA TE buffer (10mM Tris, 0.1mM EDTA pH 8.0). Single ChIP enriched DNA or control ChIP DNA should be submitted in nuclease-free H2O. Submit either source DNA as follows:

  • gDNA-Seq - a concentration between 250 - 500 ng/ul, 2-3ug total
  • ChIP-Seq - 10 ng Single ChIP enriched DNA in 30ul nuclease-free H2O

Submit samples in 1.5 -1.7ml microcentrifuge tubes with the name clearly labeled on the top of the tube with permanent marker as it is written on the order form. Please limit sample names to eight characters.

Submission of Investigator Prepared Libraries

Please follow the listed conditions:

1. Libraries must be prepared using the TruSeq Library prepartion kits or other Illumina kits compatible with the TruSeq SBS sequencing chemistry. For custom library constructs contact the GSF before submission.

2. Investigator prepared libraries must be submitted at 4nM concentration. Download the following conversion tool for converting ng/ul to nM: nM Conversion Calculator

3. When multiplexing, indiviual library concentrations must be adjusted to 4nM prior to pooling.

4. Accurate quantitation of library concentartion is critical for flow cell clustering. It is strongly recommended that qPCR be used for library quantitation.

5. It is recommended that an Agilent Bioanalyzer 2100 quality control run be performed on the library preparations to ensure proper sizing of library.

6. Please provide 15 ul - 20 ul of 4nM libraries in 1.5 -1.7ml microcentrifuge tubes with the name clearly labeled on the top of the tube with permanent marker as it is written on the order form. Please limit sample names to eight characters. 

 

Sample Submission

Hand delivery of samples can be made between 9:00 am - 4:30 pm Monday - Friday at the GSF located in RM# 1015 Hemenway, University of Kansas Medical Center. Dry ice shipments of samples, including completed order form, may be shipped overnight to:

Genomics Core
1015 Hemenway
 University of Kansas Medical Center
Receiving Dock
2106 Olathe Blvd.
Kansas City, Kansas 66160

 

For further information on the Genome Sequencing Facility browse the rest of our web site. Please feel free to call if you have any questions or require directions to the facility.

Phone: (913) 588-7127
Fax: (913) 588-7131

Last modified: Mar 10, 2014
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