Tracie Jenkins

The genetic diversity of the genus Crotalaria is unknown even though many species in this genus are economically valuable. We report the first study in which polymorphic expressed sequence tag-simple sequence repeat (EST-SSR) markers derived from Medicago and soybean were used to assess the genetic diversity of the Crotalaria germplasm collection. This collection consisted of 26 accessions representing 4 morphologically characterized species. Phylogenetic analysis partitioned accessions into 4 main groups generally along species lines and revealed that 2 accessions were incorrectly identified as Crotalaria juncea and Crotalaria spectabilis instead of Crotalaria retusa. Morphological re-examination confirmed that these 2 accessions were misclassified during curation or conservation and were indeed C. retusa. Some amplicons from Crotalaria were sequenced and their sequences showed a high similarity (89% sequence identity) to Medicago truncatula from which the EST-SSR primers were designed; however, the SSRs were completely deleted in Crotalaria. Highly distinguishing markers or more sequences are required to further classify accessions within C. juncea.

ABSTRACT Spigelia marilandica (L.) L., an herbaceous perennial native to the Southeastern United States, is used as an ornamental and for pharmacological purposes. Plant conservationists and breeders, therefore, are interested in determining its population genetic diversity. Amplified fragment length polymorphism (ALFP) fingerprints, which require upwards of 100ng of DNA, can effectively measure this diversity across the species’ range. Since DNA concentration is linked to AFLP results, an optimum protocol was developed to extract DNA from leaf material of wild-growing plants. Young leaf material was collected from 10 populations throughout Georgia, Florida, and South Carolina and stored at -80°C prior to DNA extraction. A modified Omega Bio-Tek protocol was used to extract DNA. A second protocol, which added 0.9 µL carrier RNA in the column step, was used in an attempt to increase DNA yields. Yields of DNA with and without the addition of carrier RNA were checked for quantity and quality using a 1.5% agarose gel. The majority of DNA extracted from collections which used the addition of carrier RNA showed some variation as follows. The concentration of DNA was increased from 66.6% to 830% when carrier RNA was added to samples collected in Dalton, GA. DNA extraction from 30 collections in Kathleen, GA showed variability. Of the 30 individuals collected, 13 had visible DNA only using carrier RNA, three did not generate DNA using either protocol, and of those plants that generated DNA without carrier RNA, the addition of carrier RNA increased the quantity of DNA from 40% to 500%. Additionally, a second Kathleen, GA population only produced DNA when the carrier RNA treatment was used. No DNA concentration difference resulted in extractions with or without carrier RNA from populations in Lithonia, GA and Marianna, FL. A Wakulla Springs, FL population had similar DNA quantities in 23 of 30 samples, although 7 of 30 samples collected only generated DNA when carrier RNA was used. Of 25 collections from Bonaire, GA, 19 samples resulted in DNA only when carrier RNA was used. While the remaining four samples did not produce DNA using either treatment, two produced DNA without the carrier RNA. The carrier RNA-modified Omega Bio-Tek protocol has since been effective in extracting DNA from Spigelia gentianoides, an endangered congener of S. marilandica, collected in Sneads, FL. The use of carrier RNA with the Omega Bio-Tek protocol was generally effective in getting or increasing DNA concentration in Spigelia species.

Specimens of Megacopta cribraria (Fabricius) were collected in northern Georgia in late October 2009, where they were invading homes in large numbers. This is the first known occurrence of this species and the family Plataspidae in the New World. Megacopta cribraria was previously known from Asia and Australia. A key is provided to separate Plataspidae from other families of Pentatomoidea in America North of Mexico. A diagnosis and figures are provided to facilitate recognition of M. cribraria. Reported host plants and other aspects of the biology of this species are reviewed. Megacopta cribraria is considered a pest of numerous legumes in Asia, has the potential to provide biological control of kudzu, Pueraria montana var. lobata (Willd.) Ohwi, (Fabaceae) and likely will continue to be a household pest in the vicinity of kudzu fields as well as become a pest of North American legume crops.