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Submission requirements for Purified Plasmids & PCR
Products
Template preparation
Template
DNA should be column or membrane purified using commercially
available kits such as those manufactured by Qiagen, Promega,
Millipore, etc. Elute
DNA in nuclease-free water or 10mM Tris-Cl, pH 8.0; avoid
resuspending in Tris-EDTA.
Submission format
All
templates & primers should be submitted in 96-well,
skirted PCR reaction plates (127.75mm x 85.65mm x 15mm;
Island Scientific, cat# IS 800; MJ Research, cat# MSP 9601;
Fisher Scientific, cat# 05-500-68 or VWR, cat# 53503-283). Submit
DNA and primers in separate plates, making sure that they
are arranged in the identical corresponding positions. (For
example, template DNA for sample 8 should be loaded in
well 8 of the Template Plate and its sequencing primer
is loaded in well 8 of the Primer Plate). Partially filled
plates will be charged as full plates.
Load samples in the following format :
Sample concentrations and volumes
DNA type |
Template Concentration |
Amount required/well |
Plasmid
(double-stranded) |
200 ng/uL |
10 uL |
PCR products
(<500bp)
|
5 ng/uL |
10 uL |
PCR products
(500-1000bp) |
10 ng/uL |
10 uL |
PCR products
(>1000bp) |
20 ng/uL |
10 uL |
Important Tip
Don't assume that PCR works evenly across
all wells of a plate. Make sure you quantify your product. Poor
quantification and template prep will compromise data quality.
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