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Sanger Sequencing Sample Requirements

The RTSF Genomics Core can provide short turnaround time on small and high-throughput projects utilizing the 96 capillary electrophoretic ABI 3730xl platform. The expected sequence length of PGEM plasmid controls typically ranges from 800 to 900 bp.

photo of well plates

Service Options

  • Custom (Individual Tubes, <= 16 samples)
  • High-Throughput (8-strip(s) for 8-16 samples, 96-well plate(s) for >16 samples)

Types of Reactions

  • Standard Reaction
  • 1st Tier Reaction - for high GC content and plasmids > 10kb
  • Genomic Reaction - for genomic/chromosomal DNA
  • GTP Reaction - for siRNA hairpins and secondary structure

Sample Requirements

  • The volumes requested below are sufficient for two reactions.  If less volume is provided, samples cannot be resequenced if the initial reaction fails.
  • For PCR products, it is very important to purify your sample to remove any remaining primers from the reaction, unincorporated nucleotides, enzyme, buffer, and salts.  Make sure any residual ethanol is removed as well.
  • Sample concentrations should be measured by fluorometric method (such as Qubit or PicoGreen).  UV-based measurements (such as NanoDrop) underestimate sample concentrations and should not be used.
  • Samples should be submitted in sterile water.
  • Add only one primer to each sample.  If you would like to sequence with the forward and reverse primer, two reactions are necessary.

For samples with client-added primer:

DNA Type

DNA Total Mass (ng)

Primer to add (picomoles) Total Volume (ul)
Plasmids
single-stranded DNA 200 30 12
double-stranded DNA (up to 10 kb) 1000 30 12
Purified PCR Product
<100 - 200 bp 2 - 6 30 12
200 - 500 bp 6 - 15 30 12
500 - 1000 bp 10 - 40 30 12
1000 - 2000 bp 20 - 80 30 12
>2000 bp 80 - 200 30 12

 

For samples requesting the addition of a Genomics Core standard primer:

DNA Type

DNA Total Mass (ng)

Primer to add (picomoles) Total Volume (ul)
Plasmids
single-stranded DNA 200 0 8
double-stranded DNA (up to 10 kb) 1000 0 8
Purified PCR Product
<100 - 200 bp 2 - 6 0 8
200 - 500 bp 6 - 15 0 8
500 - 1000 bp 10 - 40 0 8
1000 - 2000 bp 20 - 80 0 8
>2000 bp 80 - 200 0 8

 

Sample Submission Overview

  • We accept purified plasmids and purified PCR products.  All samples must be submitted with a completed submission form.  Submission forms can be downloaded from our Forms Webpage.
  • Sample requirements are provided in the tables above.
  • Six standard primers are offered by the Genomics Core at no additional charge.  If you will not use one of the standard primers, the customer must add their own primer to each sample.
  • For partial plate submissions, please arrange samples by column (i.e., A1-H1, A2-H2, etc.), NOT by row. A $20 re-racking fee will be charged for any samples not arranged in columns.
  • Sequences can be accessed and downloaded from the Genomics Depot. If your data is not available after 48 hours please contact us at gtsf@msu.edu.  Data is only available for 6 months.  See our Data Retention Policy webpage for more information.

Sample Drop-off/Shipping samples

There are three options for submitting samples:

  1. Samples can be dropped off at the RTSF Genomics Core submission freezer, which is outside of room S18 in the Plant Biology building (basement of the south wing).
  2. Samples can be dropped off at our drop box located across the street on the 5th floor of the BPS building. Samples are picked up at 8:30 AM and processed that day.
  3. Samples can be shipped to the Genomics Core.  Please ship Sanger sequencing samples to the following address:

    Colleen Curry
    612 Wilson Road, Rm S-18
    East Lansing, MI 48824