{"id":{"repo_id":"wku-diss","oai_identifier":"oai:digitalcommons.wku.edu:theses-1670"},"canonical_url":"https://search.dev.ndltd.org/etd/wku-diss/oai:digitalcommons.wku.edu:theses-1670","repository":{"repo_id":"wku-diss","name":"Western Kentucky University","base_url":"https://digitalcommons.wku.edu/do/oai/"},"display":{"title":"Secondary Level Screening of Chlamydomonas Reinhardtii Mutants Defective in Circadian Gene Expression","abstract":"To elucidate the signal transduction chain mediating circadian clock control, this work focuses on the isolation of Chlamydomonas reinhardtii mutants which are defective in circadian gene expression. In a previous study, the reporter gene ARS2 encoding the arylsulfatase enzyme was fused to the promoter of the circadian-regulated CABII-1 gene and transformed into the Chlamydomonas nucleus. The ble marker was introduced into the genome of this transformant via insertional mutagenesis to generate mutants defective in circadian CABII-1 expression. Potential mutants were selected based on aberrant single-point accumulative arylsulfatase activity. In this study, the arylsulfatase activity over the entire growth cycle was further investigated in these mutants and the reliability of the single-point screen was assessed. Of the 16 strains whose accumulative arylsulfatase activity did not differ from the nonmutagenized control in the single-point screen, 12 still showed no significant difference in a multiple-point screen. Of the 9 potential mutants with significant difference to the control in the single-point screen, 3 showed no significant difference in the multiple-point screen. Subsequently, 8 of the candidate mutants with aberrant reporter enzyme activity in the multiple-point screen were characterized by the abundance of their mRNA. The peak-to-trough ratio of CABII-1 and ARS2 transcript abundance was significantly reduced in 4 of these mutants.","abstract_html":"To elucidate the signal transduction chain mediating circadian clock control, this work focuses on the isolation of Chlamydomonas reinhardtii mutants which are defective in circadian gene expression. In a previous study, the reporter gene ARS2 encoding the arylsulfatase enzyme was fused to the promoter of the circadian-regulated CABII-1 gene and transformed into the Chlamydomonas nucleus. The ble marker was introduced into the genome of this transformant via insertional mutagenesis to generate mutants defective in circadian CABII-1 expression. Potential mutants were selected based on aberrant single-point accumulative arylsulfatase activity. In this study, the arylsulfatase activity over the entire growth cycle was further investigated in these mutants and the reliability of the single-point screen was assessed. Of the 16 strains whose accumulative arylsulfatase activity did not differ from the nonmutagenized control in the single-point screen, 12 still showed no significant difference in a multiple-point screen. Of the 9 potential mutants with significant difference to the control in the single-point screen, 3 showed no significant difference in the multiple-point screen. Subsequently, 8 of the candidate mutants with aberrant reporter enzyme activity in the multiple-point screen were characterized by the abundance of their mRNA. The peak-to-trough ratio of CABII-1 and ARS2 transcript abundance was significantly reduced in 4 of these mutants.","abstract_has_math":false,"creators":["Huang, Mingya"],"institution":null,"degree_name":"Master of Science","degree_level":null,"degree_discipline":"Department of Biology","degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2001,"date_issued":"2001-08-01T07:00:00Z","date_published":"2001-08-01T07:00:00Z","updated_at":"2026-07-24T06:07:42Z","subjects":["Medical Sciences"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://digitalcommons.wku.edu/theses/667","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["Huang, Mingya"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Department of Biology"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Master of Science"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Medical Sciences"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://digitalcommons.wku.edu/theses/667"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["To elucidate the signal transduction chain mediating circadian clock control, this work focuses on the isolation of Chlamydomonas reinhardtii mutants which are defective in circadian gene expression. In a previous study, the reporter gene ARS2 encoding the arylsulfatase enzyme was fused to the promoter of the circadian-regulated CABII-1 gene and transformed into the Chlamydomonas nucleus. The ble marker was introduced into the genome of this transformant via insertional mutagenesis to generate mutants defective in circadian CABII-1 expression. Potential mutants were selected based on aberrant single-point accumulative arylsulfatase activity. In this study, the arylsulfatase activity over the entire growth cycle was further investigated in these mutants and the reliability of the single-point screen was assessed. Of the 16 strains whose accumulative arylsulfatase activity did not differ from the nonmutagenized control in the single-point screen, 12 still showed no significant difference in a multiple-point screen. Of the 9 potential mutants with significant difference to the control in the single-point screen, 3 showed no significant difference in the multiple-point screen. Subsequently, 8 of the candidate mutants with aberrant reporter enzyme activity in the multiple-point screen were characterized by the abundance of their mRNA. The peak-to-trough ratio of CABII-1 and ARS2 transcript abundance was significantly reduced in 4 of these mutants."]},{"key":"dc:title","label":"Title","values":["Secondary Level Screening of Chlamydomonas Reinhardtii Mutants Defective in Circadian Gene Expression"]}]}],"canonical_facts":{"dc:creator":["Huang, Mingya"],"dc:description.abstract":["To elucidate the signal transduction chain mediating circadian clock control, this work focuses on the isolation of Chlamydomonas reinhardtii mutants which are defective in circadian gene expression. In a previous study, the reporter gene ARS2 encoding the arylsulfatase enzyme was fused to the promoter of the circadian-regulated CABII-1 gene and transformed into the Chlamydomonas nucleus. The ble marker was introduced into the genome of this transformant via insertional mutagenesis to generate mutants defective in circadian CABII-1 expression. Potential mutants were selected based on aberrant single-point accumulative arylsulfatase activity. In this study, the arylsulfatase activity over the entire growth cycle was further investigated in these mutants and the reliability of the single-point screen was assessed. Of the 16 strains whose accumulative arylsulfatase activity did not differ from the nonmutagenized control in the single-point screen, 12 still showed no significant difference in a multiple-point screen. Of the 9 potential mutants with significant difference to the control in the single-point screen, 3 showed no significant difference in the multiple-point screen. Subsequently, 8 of the candidate mutants with aberrant reporter enzyme activity in the multiple-point screen were characterized by the abundance of their mRNA. The peak-to-trough ratio of CABII-1 and ARS2 transcript abundance was significantly reduced in 4 of these mutants."],"dc:identifier":["https://digitalcommons.wku.edu/theses/667"],"dc:subject":["Medical Sciences"],"dc:title":["Secondary Level Screening of Chlamydomonas Reinhardtii Mutants Defective in Circadian Gene Expression"],"dc:type":["Thesis"],"thesis:degree_discipline":["Department of Biology"],"thesis:degree_name":["Master of Science"]},"updated_at":"2026-07-24T06:07:42Z"}