Services - DNA Sequencing

A Brief Guide to Interpreting DNA Sequencing Electropherogram (Version 3.0)

This guide includes an example of high quality sequence as well as many different problems that occur with DNA sequences attained from the 3730 DNA Analyzer in the Plant-Microbe Genomics Facility. For those figures that demonstrate a problem there is a (1) description of the problem, (2) the most likely cause(s), and (3) one or more solutions.

Definition of term Causes of the Problem
700 or more high quality base assignments -No problem
less than 700 high quality base assignments -low quality or quantity of template or primer
-primer has poor binding
-high G/C content in template
-strong or weak stop (see Fig. 9)
peaks under peaks which are in phase and are reported as "Ns" -multiple templates or priming sites (see Fig. 7)
-primer N minus 1 (see Fig. 12)
-homoN slippage (see Fig. 8)
sequence is 5 "Ns" -lack of extension products (see Fig. 6)
loss of peak resolution by progressive peak widening -sample contamination (see Fig. 11)

If you have additional questions or comments about the figures below, then please do not hesitate to contact the facility.

Fig. 1 - Electropherogram with high quality sequence Fig. 2 - Electropherogram demonstrating Quality Values assigned to each base by Sequencing Analysis ver 5.1 Fig. 3 - Electropherogram that demonstrates the limit of the resolution of the 3730 DNA Analyzer
Fig. 1
Electropherogram with
high quality sequence
Fig. 2
Electropherogram demonstrating Quality Values assigned to each base by Sequencing Analysis ver 5.1
Fig. 3
Electropherogram that demonstrates the limit of the resolution of the 3730 DNA Analyzer
Fig. 4 - Electropherogram with unincorporated nucleotide peaks Fig. 5 - Electropherogram with mobility errors

Fig. 6a - Electropherogram that demonstrates a lack of extension products, i. e. no bands

Fig. 6b - Electropherogram that demonstrates a lack of extension products,i. e. no bands
Fig. 4
Electropherogram with unincorporated nucleotide peaks 		Fig. 5
Electropherogram with mobility errors 		Figs. 6a,b
Electropherogram that demonstrates
a lack of extension products,
i. e. no bands

Fig. 7 - Electropherogram that has multiple sequences

Fig. 7b - Electropherogram that has multiple sequences

Fig. 8a - Electropherograms that have homoN slippage

Fig. 8b - Electropherograms that have homoN slippage

 Fig. 9 - Electropherogram that has a strong stop
Fig. 7a,b
Electropherogram that has
multiple sequences 		Fig. 8a,b
Electropherograms that have
homoN slippage 		Fig. 9
Electropherogram that has a strong stop
Fig. 10 - Electropherogram that has a micro-air bubble or debris

Fig. 11a - Electropherogram that has 'the spread'

Fig. 11b - Electropherogram that has 'the spread'

 Fig. 12 - Electropherogram that has Primer N-1
Fig. 10
Electropherogram that has a micro-air bubble or debris 		FIg. 11a,b
Electropherogram that has "the spread"		 Fig. 12
Electropherogram that has Primer N-1