This project was initially made
possible
with support from the Governor's 2010 Initiative.
Currently this project is supported by a grant from the South Dakota
Agricultural Experiment Station
| Cuphea is a genus of
herbaceous plants originating in the Western hemisphere. Several
species of Cuphea were found to be a rich source of medium-chain fatty
acids, which are important for a number of nutritional and industrial
uses. These oils are currently available only from tropical oils,
which must be imported into the United States. Efforts are
underway to domesticate this plant, but because Cuphea has only
recently
been subjected to artificial selection, it has persistent attributes
that limit its agronomic potential. Three most notable problems
are seed dormancy, seed shattering and indeterminate flowering. |
Cuphea |
I
am collaborating with a small group of investigators from SDSU and the
USDA-ARS in Morris, MN to study this plant and assist in its
domestication. Work on this plant was resently featured in the SDSU
publication
Farm and Home Research. (To left: Abdullah Jaradat, USDA-ARS
Morris,MN and Jose Gonzalez, SDSU) 
Cuphea plant
Marker discovery: My main objective of this project is to find DNA markers, specifically SSRs, that can be used to assist breeders in the selection of desirable traits. SSR stands for “simple sequence repeat." These are two to four base sequences that are repeated several consecutive times in DNA. SSRs, also called microsatellites, are often polymorphic within a population for the number of repeats. Here’s an example of an SSR involving a trinucleotide repeat:
5’-…ATGATGATGATGATG…-3’
Our immediate goal is to find sufficient SSRs to saturate the Cuphea genome. Correlations can then be made between SSRs and beneficial phenotypic traits for domesticated Cuphea.
Our strategy for SSR
discovery follows Edwards et al.
(1996) and is roughly as follows: First digest extracted Cuphea
DNA with RsaI. A blunt-end
ligation is then performed using the digested fragments and special
sequence-specified adaptor segments, which contain a MluI restriction
site. The resulting DNA strands are denatured with heat and
hybridized to a positively charged nylon filter embedded with
dinucleotide repeats. The filter is thoroughly washed to remove
the Cuphea DNA that does not contain segments complimentary to the
dinucleotides on the filter. The hybridized DNA is then amplified
using PCR with added adaptor segments serving as the primer.
Amplified DNA and plasmids are then digested with MluI, ligated and used
to transform bacteria for cloning. Bacteria containing plasmids
with our insert will be identified using blue/white selection and their
plasmids will be isolated for sequencing. (Library and gel made by Nick Gau,
undergraduate student
researcher.) Edwards, K.J., J.H.A. Barker, A. Daly, C. Jones and A. Karp 1996. Microsatellite libraries enriched for several microsatellite sequences in plants. BioTechniques 20:758-760.