- Lee, Jung-Lim
- Delaware State University
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This proposal is an integration project for Teaching, Outreach, and Research. The research of this proposed project will be accomplished by a multidisciplinary team of food scientists, molecular biologists, nutritionists, and bioinformatics scientists in the academic institution, government agency, and private sector. This project will focus on the new sequencing and genomics technologies that impact food, nutrition, and agricultural research. This study will also develop an innovative curriculum toeducate and train skilled college students as well as to attract high school students to the DSU Food and Nutritional Science Program.
The specific objectives of Teaching & Outreach are: 1) To develop the multidisciplinary new course including lectures and labs and to entice new students for DSU. 2) To establish the new course by evaluation and feedback of the curriculum for continuous improvement. 3) To disseminate this project accomplishment throughout the communities and recruit students into the Food and Nutritional Science Program at DSU.
The specific objectives of the Research portion are: 1) To screen and identify bacteriocin having antimicrobial activity against spoilage bacteria.2) To investigate the antimicrobial effect of several natural preservatives including salt, lemon, grape fruit seed extract, and bacteriocin. 3) To analyze the microbial diversity of non-treated or natural preservative-treated catfish during refrigerated storage by metagenomic assay using Illumina sequencing.
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The new genomic DNA analysis has been driven by the introduction of next-generation sequencing (NGS) platforms. NGS has markedly accelerated numerous fields of biological research, enabling researchers to pursue experiments that previously were not technically affordable. Despite the promise of such technologies, there has been only limited research in foods. This project aims to provide students with the critical knowledge and skills that will empower them to participate in the next generation of food safety research. Toward this aim a new multidisciplinary course reinforcing applications of genomics for the undergraduate Food and Nutritional Science major will be developed. The new course will contain principles of molecular biology, genomics, and its applications to food science and nutrition. The course will consist of both lectures and labs. In addition, summer laboratory workshops will be developed for students at local high schools and through the 4-H program to provide deeper scientific knowledge and motivation through hands-on experience in order to attract students' interest in food science and agricultural majors. The project further enables graduate students to participate in and drive experiments applying the NGS technique to identify the bacterial species of fishery products treated with natural preservatives. Catfish will be treated with the four kinds of natural preservatives during storage in the refrigerator and bacterial diversity of the fish microflora will be determined following NGS assay. Innovative coursework and applied research aspects of this project will synergize to increase undergraduate and graduate students' knowledge and competence in cutting-edge molecular biology providing skills that can lead to new career opportunities in the food and agricultural industries. This integrated project will contribute to enhancing the safety of fishery products, reducing outbreaks of food poisoning, and strengthening the capacity of the Masters of Science Food Science Program.
Teaching 1.The lecturers of this multidisciplinary course will consist with several faculty and outside experts. The new course will focus on current food microbiology and molecular biology, genomics, next generation sequencing, and applications. This project will open a 3-credit-hour course titled ""Food Genomics"" consisting of lectures and labs. 2.In order to announce this new course, the PD and staff will develop newsletters and brochures containing the course introduction and a synopsis of the curriculum as well as will be distributed to new students at the DSU New Student Orientation, pre-major (undecided) students, and updated on the college and DSU websites. 3.Students will conduct lab experiments such as culture medium for bacterial propagation, bacteria inoculation, DNA extractions, PCR, enzyme digestion, DNA electrophoresis, DAN sequencing, etc. We will videotape lab activities and procedure for each experiment and upload on the DSU Blackboard and the PD's lab website. 4.Students will be given exams and quizzes and will be required to provide lab reports which will be evaluated to assess the students' progress. The class will also be evaluated by peer evaluators in the middle of the semester. At the end of the semester, the class will be evaluated through a survey of students in this course. Outreach 1. The special summer laboratory workshops entitled "Food Biotechnology for Food Safety & Health" will be open for the local high school, 4-H students, and their parents/teachers. 2. Faculty and staff will create newsletters, pamphlets, and brochures introducing the summer lab workshop and career opportunities in food and agriculture. These materials will be distributed to students, parents, and teachers in the local high schools. 3. Ten-day apprenticeship program will be conducted during the summer in the Department of Human Ecology at DSU. The activities will be organized Monday to Friday. Throughout the summer program, students will receive lecture and practical experience with molecular biology equipment and procedures. 4. At the end of the program, students will be expected to submit a report and make a 10 minute PowerPoint presentation on what they have learned. 5. The Department will confer certificates to the students, who complete the workshop. Faculty and 4-H leader will identify potential students who are interested in entering the Food & Nutritional Science Program and could offer them mentorship before they come to DSU. Research 1. The cultured LAB isolates will be tested by agar diffusion assay for their antimicrobial effects against fish spoilage bacteria such as Listeria, Pseudomonas, Photobacterium, and Shewanella species. 2. The screened LAB supernatants will be crudely purified by precipitation and then the antimicrobial activities of purified supernatant will be confirmed by the paper disc method. The supernatants containing antimicrobial activities will be used as bacteriocins for treatment on the fish. 3. To identify the bacteriocin producing LAB strain, 16S rRNA gene will be amplified by PCR. Target DNA sequence of 16S rRNA will be amplified with primers pairs (16SPO/16SP6) and then sequenced. The species of the LAB strains will be identified by comparing the homology with the Genbank data base. 4. The stomacher bags containing catfish fillets will be stored at 4°C for 1 hr to marinate the fillets in the natural preservatives (sea salt, lemon extract, grapefruit seed extract, and screened bacteriocin) and then excess solution will be drained out. The stomacher bags containing the fillets will be stored at 4°C for 20 days and total bacteria will be counted every 2 days. Also, sensory evaluation will be performed to evaluate the acceptances during the period at refrigerated storage. 5. The four natural preservatives will be added to catfish according to the selected concentration in the previous experiments and stored at 4°C. Every 4 days, total bacterial genomic DNA will be extracted from fillets using an optimized method and enumerated for total bacteria using the culture-method. The extracted DNA will be used in Illumina sequencing. 6. The V1-V3 (27F-534R) region of the 16S rRNA gene will be amplified using modified 27F and 534R primers. A custom series of barcodes developed by the Institute for Genome Science (University of Maryland) will be included on the reverse primer allowing multiplexing of up to 96 samples per sequencing reaction. 7. PCR products of the correct size will be recovered using a gel extraction kit. For each library, PCR products with unique indexes will be mixed in equal nanogram quantities, quantified by Qubit fluorometric method, and sequenced on an Illumina MiSeq instrument. Finally, Illumina sequenced amplicon libraries will be analyzed using the QIIME amplicon analysis pipeline.
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- Nat'l. Inst. of Food and Agriculture
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- Bacterial Pathogens