Functional studies of lignin biosynthesis genes and putative flowering gene in Miscanthus x giganteus and studies on indolyl glucosinolate biosynthesis and translocation in Brassica oleracea

The first topic area of this thesis involved studies on the accumulation and translocation of glucosinolates (GSs), bioactive secondary plant compounds, in broccoli plants. Changes in GS accumulation and gene expression levels in response to exogeneous methyl jasmonate (MeJA) treatment were analyzed in different tissue types at different developmental stages of broccoli. Greater accumulation of GSs with MeJA treatment was observed in apical leaves of broccoli seedlings and florets of plants at harvest maturity. Increases in indolyl GS in apical leaves of seedlings and florets were coupled with the up-regulation of indolyl GS biosynthesis genes. The accumulation of indolyl GSs appears to be modulated by MeJA treatment in an organ-specific manner for optimal distribution of defense substances in the plant. Metabolic profiling of hydrophilic metabolites using GC-MS demonstrated increased accumulation of various phenolics, ascorbates and amino acids in broccoli tissues after MeJA treatment. Distinct changes in carbohydrate levels observed between different tissues (vegetative leaves and floret tissues) of broccoli plants after treatment suggest that carbon metabolism is differentially modulated by MeJA treatment in different tissue types depending on sink-source relationships. Reduced levels of hexose sugars and tricarboxylic acid intermediates after MeJA treatment may reflect the increased requirement for carbon and energy needed to drive secondary product biosynthesis to accumulate metabolites for defense against insects and other herbivores. Substantial increases of indolyl and aromatic GSs after exogenous treatment with MeJA in stem and petioles of seedlings and the existence of intact indolyl-GS forms in phloem exudates suggest enhanced de novo synthesis in combination with active transport. Indoly GSs share structural similarities with the auxin, IAA, and may interact with components of the auxin transport system for intra- and extra-cellular transport or translocation. Application of the auxin efflux inhibitor, 1-naphthylphthalamic acid (NPA) reduced MeJA-mediated accumulation of indolyl GSs in broccoli florets and seedling tissues. NPA did not inhibit expression of indolyl GS biosynthesis genes shown to be upregulated by MeJA treatment or the accumulation of tryptophan, the amino acid precursor of indolyl GSs. Exogenous application of benzyl GS to Arabidopsis roots induced ectopic expression of the PIN1 protein associated with the auxin transport system similar to treatment with NPA, again suggesting GS interaction with the auxin efflux carrier system. The inhibitory effect of NPA on MeJA-mediated accumulation of GS may be due to competitive binding of NPA to auxin efflux carrier components and that GS transport is mediated by the auxin transport system. The inhibitory effect of NPA on indolyl and aromatic GS accumulation and the bioactivity of exogenous treatment of these GS compounds in PIN1 localization, Arabidopsis root growth, and gravitrophic response suggest that indolyl and aromatic GSs may be antagonistic to IAA transport and biosynthesis. Indolyl and aromatic GSs can also be potentially converted into IAA by hydrolysis. This intrinsic feature of GSs may be the part of a sophisticated regulatory process where the metabolic pathways in the plant shift from active growth to a reversible defense posture in response to biotic or abiotic stress. It seems likely that indolyl and aromatic GSs are important compounds that provide connections between jasmonate and auxin signaling. Further studies are required to reveal the regulatory mechanism for crosstalk between the two hormones. The third part of this research was to investigate effect of selenium fertilization and MeJA treatment on accumulation of GSs in broccoli florets. Increasing dietary intake of the element selenium (Se) has been shown to reduce the risk of cancer. Simultaneous enhancement of both Se and GS concentrations in broccoli floret tissue were conducted through the combined treatment of MeJA with Se fertilization. A low level of Se fertilization (concentration) with MeJA treatment displayed no significant changes in total aliphatic GS concentrations with 90% and 50% increases in indolyl and total GSs concentrations, respectively. This result suggests that Se- and GS-enriched broccoli with improved health-promoting properties can be generated by this combined treatment. The second topic of this thesis was conducted to provide basic information required to improve biomass quality and productivity and develop tools for gene transformation in Miscanthus x giganteus. The perennial rhizomatous grass, Miscanthus x giganteus is an ideal biomass crop due to its rapid vegetative growth and high biomass yield potential. As a naturally occurring sterile hybrid, M. x giganteus must be propagated vegetatively by mechanicalling divided rhizomes or from micropropagated plantlets. The effect of callus type, age and culture methods on regeneration competence was studied to improve regeneration efficiency and shorten the period of tissue culture in M. x giganteus propagation. Seven lignin biosynthesis genes and one putative flowering gene were isolated from M. x giganteus by PCR reactions using maize othologous sequences. Southern hybridization and nuclear DNA content analysis indicated that the genes isolated from M. x giganteus exist in the genome of other Miscanthus species as multiple copies. Analysis of lignin content and histological staining of lignin deposition indicated that higher lignin content is found in mature stem node tissues compared to young leaves and apical stem nodal tissues. Cell wall lignification is associated with increasing tissue maturity in Miscanthus species. RNAi and antisense constructs harboring sequences of these genes were developed to generate Miscanthus transgenic plants with suppressed of lignin biosynthesis and delayed flowering.

English syllable confusion and imitation in Korean bilingual and monolingual children and adults

In the present study, Korean-English bilingual (KEB) and Korean monolingual (KM) children, between the ages of 8 and 13 years, and KEB adults, ages 18 and older, were examined with one speech perception task, called the Nonsense Syllable Confusion Matrix (NSCM) task (Allen, 2005), and two production tasks, called the Nonsense Syllable Imitation Task (NSIT) and the Nonword Repetition Task (NRT; Dollaghan & Campbell, 1998). The present study examined (a) which English sounds on the NSCM task were identified less well, presumably due to interference from Korean phonology, in bilinguals learning English as a second language (L2) and in monolinguals learning English as a foreign language (FL); (b) which English phonemes on the NSIT were more challenging for bilinguals and monolinguals to produce; (c) whether perception on the NSCM task is related to production on the NSIT, or phonological awareness, as measured by the NRT; and (d) whether perception and production differ in three age-language status groups (i.e., KEB children, KEB adults, and KM children) and in three proficiency subgroups of KEB children (i.e., English-dominant, ED; balanced, BAL; and Korean-dominant, KD). In order to determine English proficiency in each group, language samples were extensively and rigorously analyzed, using software, called Systematic Analysis of Language Transcripts (SALT). Length of samples in complete and intelligible utterances, number of different and total words (NDW and NTW, respectively), speech rate in words per minute (WPM), and number of grammatical errors, mazes, and abandoned utterances were measured and compared among the three initial groups and the three proficiency subgroups. Results of the language sample analysis (LSA) showed significant group differences only between the KEBs and the KM children, but not between the KEB children and adults. Nonetheless, compared to normative means (from a sample length- and age-matched database provided by SALT), the KEB adult group and the KD subgroup produced English at significantly slower speech rates than expected for monolingual, English-speaking counterparts. Two existing models of bilingual speech perception and production—the Speech Learning Model or SLM (Flege, 1987, 1992) and the Perceptual Assimilation Model or PAM (Best, McRoberts, & Sithole, 1988; Best, McRoberts, & Goodell, 2001)—were considered to see if they could account for the perceptual and production patterns evident in the present study. The selected English sounds for stimuli in the NSCM task and the NSIT were 10 consonants, /p, b, k, g, f, θ, s, z, ʧ, ʤ/, and 3 vowels /I, ɛ, æ/, which were used to create 30 nonsense syllables in a consonant-vowel structure. Based on phonetic or phonemic differences between the two languages, English sounds were categorized either as familiar sounds—namely, English sounds that are similar, but not identical, to L1 Korean, including /p, k, s, ʧ, ɛ/—or unfamiliar sounds—namely, English sounds that are new to L1, including /b, g, f, θ, z, ʤ, I, æ/. The results of the NSCM task showed that (a) consonants were perceived correctly more often than vowels, (b) familiar sounds were perceived correctly more often than unfamiliar ones, and (c) familiar consonants were perceived correctly more often than unfamiliar ones across the three age-language status groups and across the three proficiency subgroups; and (d) the KEB children perceived correctly more often than the KEB adults, the KEB children and adults perceived correctly more often than the KM children, and the ED and BAL subgroups perceived correctly more often than the KD subgroup. The results of the NSIT showed (a) consonants were produced more accurately than vowels, and (b) familiar sounds were produced more accurately than unfamiliar ones, across the three age-language status groups. Also, (c) familiar consonants were produced more accurately than unfamiliar ones in the KEB and KM child groups, and (d) unfamiliar vowels were produced more accurately than a familiar one in the KEB child group, but the reverse was true in the KEB adult and KM child groups. The KEB children produced sounds correctly significantly more often than the KM children and the KEB adults, though the percent correct differences were smaller than for perception. Production differences were not found among the three proficiency subgroups. Perception on the NSCM task was compared to production on the NSIT and NRT. Weak positive correlations were found between perception and production (NSIT) for unfamiliar consonants and sounds, whereas a weak negative correlation was found for unfamiliar vowels. Several correlations were significant for perceptual performance on the NSCM task and overall production performance on the NRT: for unfamiliar consonants, unfamiliar vowels, unfamiliar sounds, consonants, vowels, and overall performance on the NSCM task. Nonetheless, no significant correlation was found between production on the NSIT and NRT. Evidently these are two very different production tasks, where immediate imitation of single syllables on the NSIT results in high performance for all groups. Findings of the present study suggest that (a) perception and production of L2 consonants differ from those of vowels; (b) perception and production of L2 sounds involve an interaction of sound type and familiarity; (c) a weak relation exists between perception and production performance for unfamiliar sounds; and (d) L2 experience generally predicts perceptual and production performance. The present study yields several conclusions. The first is that familiarity of sounds is an important influence on L2 learning, as claimed by both SLM and PAM. In the present study, familiar sounds were perceived and produced correctly more often than unfamiliar ones in most cases, in keeping with PAM, though experienced L2 learners (i.e., the KEB children) produced unfamiliar vowels better than familiar ones, in keeping with SLM. Nonetheless, the second conclusion is that neither SLM nor PAM consistently and thoroughly explains the results of the present study. This is because both theories assume that the influence of L1 on the perception of L2 consonants and vowels works in the same way as for production of them. The third and fourth conclusions are two proposed arguments: that perception and production of consonants are different than for vowels, and that sound type interacts with familiarity and L2 experience. These two arguments can best explain the current findings. These findings may help us to develop educational curricula for bilingual individuals listening to and articulating English. Further, the extensive analysis of spontaneous speech in the present study should contribute to the specification of parameters for normal language development and function in Korean-English bilingual children and adults.