AGRICULTURAL SCIENCES
A Comparison of Agarose, Metaphor Agarose, and Polyacrylamide Gel Electrophoresis Systems in
Resolving Pawpaw Simple Sequence Repeat Markers
LAUREN A. COLLINS*, JEREMIAH D. LOWE, and KIRK W. POMPER. Land Grant Program,
Kentucky State University, Atwood Research Facility, Frankfort, KY 40601-2355
Pawpaw [Asimina triloba (L.) Dunal] is a tree fruit native to much of the Eastern United States. Since 1994,
Kentucky State University (KSU) has served as the USDA National Clonal Germplasm Repository, or gene
bank, for pawpaw; therefore, the assessment of genetic diversity with molecular markers in pawpaw is an
important research priority for the KSU program. Using the polymerase chain reaction (PCR) with simple
sequence repeat (SSR) primers and pawpaw template DNA, products between 150 to 500 bp are usually
amplified and visualized via electorphoresis. The objective of this study was to determine if SSR-PCR
products separated on agarose, metaphor agarose, and polyacrylamide gel electrophoresis systems display
unique scoring patterns that result in different genetic separation upon analysis for nine pawpaw cultivars.
DNA was extracted from young leaves collected from the pawpaw cultivars: Cales Creek, Davis,
Middletown, NC-1, Overleese, Rebeccas Gold, Taytwo, Wilson, and Zimmerman using a DNAmite kit.
SSRs primers B129 and C104 were developed by Genetic Information Services (Chatsworth, CA) and were
used to amplify the pawpaw template DNA. PCR products were separated by electrophoresis on 3%
agarose, 3% metaphor agarose, and 5% polyacrylamide gels. Following electrophoresis, gels were stained
with ethidium bromide and photographed. Gel analysis was carried out using Kodak 1D software.
Dendrograms were produced from the marker scoring data using NTSYSpc v2.11T. Each electrophoresis
system produced a unique separation of the pawpaw cultivars, although some groupings were similar using
the different systems.