Document Type
Thesis - University Access Only
Award Date
2004
Degree Name
Master of Science (MS)
Department / School
Plant Science
First Advisor
Fedora Sutton
Abstract
Potassium ion channels play a vital role in transporting K + across the cell membrane of higher plants. The activities of potassium ion channels are regulated by endogenous and environmental factors. Previous experiments have already proven that potassium ion channels are components in the ABA signaling pathway. In this study, we have been trying to identify, clone, and characterize the outward K + channel from Vicia Jaba guard cell in order to investigate the ABA signaling transduction pathway, and finally to utilize that information to isolate the ABA receptor. The requested cloned guard cell outward rectifying K + channel (GORK) cDNA (Ache et al., 2000) was never sent. Therefore, we undertook the isolation of the Vicia Jaba GORK. One cDNA clone was obtained from the total RNA of Vicia Jaba guard cells by RT-PCR (reverse transcription followed by polymerase chain reaction) with Arabidopsis guard cell outward K + channel specific primers. The amplicon produced was of a size similar to that obtained with the positive control Arabidopsis guard cell RNA (approximately 500bp ). This amplicon was inserted in the vector pCR2. l-TOPO, and sequenced. A database search showed that this fragment lacks homology with any K + channel. The sequence of this fragment matched that of a dehydrogenase gene. Arabidopsis genomic DNA was used to test the ability of the primers to amplify the GORK segment. A product of approximate 2Kb was obtained. The sequence of this product matched GORK gene. This segment was used as the probe in both northern and southern hybridization analysis of Vicia faba. No product was detected. The reason for lack of homology with Arabidopsis guard cell outward K + gene is under investigation. In order to check whether Vicia faba guard cell mRNA expresses an outward rectifying K + channel in Xenopus laevis oocytes, the two-electrode voltage clamp (TEVC) technique was used to monitor the oocytes injected with Viciafaba guard cell mRNA. A small outward current (1200nA at 160mV test potential) was produced. This outward current was not reduced significantly by er channel blocker (9-AC). This result suggests that Viciafaba guard cell mRNA-injected oocytes express the outward current.
Library of Congress Subject Headings
Fava bean -- Genetics.
Potassium channels.
Potassium -- Physiological transport.
Cellular signal transduction.
Publisher
South Dakota State University
Recommended Citation
He, Dong, "Isolation and Identification of the Outward K+ Channel Genes From Vicia Faba Guard Cell" (2004). Electronic Theses and Dissertations. 1068.
https://openprairie.sdstate.edu/etd2/1068
Rights Statement
