Identifying zero sequence diversity genes in maize using Temperature Gradient Capillary Electrophoresis (TGCE)

1
Identifying zero sequence diversity genes in
maize using Temperature Gradient Capillary
Electrophoresis (TGCE)
Julie M. Meyer1, Hsin Debbie Chen2, Josh
Shendelman2, Patrick S. Schnable2
1 Truman State University, Kirksville, MO
63501 2 Iowa State University, Ames, IA 50011
Results

• Of the 81 primer pairs retested, data was
  obtained from 67. 66 were confirmed to have zero
  sequence diversity and one failed to be
  confirmed.
• Multiple peaks indicate that more than one
  sequence of DNA is present while single peaks
  indicate the presence of only one DNA sequence.

Twenty-four inbred lines of maize were tested
for zero sequence diversity genes using primers
designed from Maize Assembled Genomic Islands
(MAGI) sequences. After no polymorphisms were
observed on an agarose gel, they were run on TGCE
instrument to detect the presence of
polymorphisms. Data was obtained for 67 primers
and 66 were confirmed to have zero sequence
diversity. Further testing of teosinte lines will
show which of the zero sequence diversity genes
in maize are responsible for the domestic
phenotype of modern maize.

• Genes that exhibit no diversity might be the key
  genes responsible for the domestic phenotype of
  maize compared to its wild relatives.
• TGCE is a faster, cheaper and more accurate way
  of determining if two sequences are in fact
  distinct.
• Heteroduplexes form when two distinct strands of
  DNA anneal together. These are displayed on a
  graph as multiple peaks. See Figure 1.
• Homoduplexes form when two identical strands of
  DNA anneal together. These are displayed on a
  graph as a single peak.

Figure 1. Top shows a single peak. Bottom shows
multiple peaks.

• Further testing of these primers on teosinte
  inbred lines is needed to identify genes
  responsible for maize domestication traits.
• Those genes with high levels of diversity in
  teosinte but zero diversity in maize could be
  genes that influenced the domestication of maize.
  
• It is important to understand these genes because
  they are the traits important to humans and they
  might shed more light of the process of
  domestication.

Methods

• Twenty-four maize inbred lines tested.
• 81 primers designed from MAGI sequences.
• After PCR, run on 1 agarose gel to find
  polymorphisms. See Figure 2.
• If none, DNA was combined with B73 DNA and
  allowed to denature and reanneal.
• Run on TGCE to find polymorphisms.
• All primers tested at least twice to confirm.

Figure 2. Agarose gel electrophoresis. Top
shows a polymorphic gene. Bottom shows a
candidate zero diversity gene.
This project was funded by the National Science
Foundation grant (DBI-0321711).