Progress 10/01/12 to 09/30/13
Outputs
Target Audience: During the initial project period the target audience included graduate and undergraduate students learning sequencing,gene identification, and lab techniques andskills. Changes/Problems: Project director accepted aposition at the University of Wisconsinand terminated this UCONN Hatch Multistate project in April, 2013. What opportunities for training and professional development has the project provided? Opportunities for training included learning new techniques and skills by both the graduate and undergraduate trainees and the PI. Students were taught sample collection and processing, and data analysis. How have the results been disseminated to communities of interest? The PI and one undergraduate student attended the Joint Annual meetings, (ASAS, ADSA) and presented research findings. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Objective 1. Dairy cattle experiments were conducted tocollect liver and blood samples from dry and lactating cattle. Sequencing technology was used to sequence and identify SNP variations within target genes.An antibody was validated and traditional Western Blot analysis was conducted to allow for validation of the PCR based protein assay. Objective 2. Executionof a dairy cattleexperimentincluded collection of liver and blood samples from dry and lactaing cattle, healthy or diagnosed ketosis, from the University of Connecticut dairy herd. The samples were used forsequencing, protein analysis, and liver lipid quantification (n=16). Fundamental knowledge was gained regarding target genes and SNP presence. In addition, specific fundamental knowledge was gainedin relation toanovel target gene, bovine adiponutirn. Expression of mRNA and protein abundance reveal that this gene is regulated during the transition to lactation period in dairy cattle. Sequencing and SNP identificaton of this gene wasperformed. Five undergraduate and twomasters(one non-thesis and one thesis) students were mentored by the PI. These students were taught sample collection and processing techniques and data analysis.
Publications
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Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: The objectives of this PI as a part of the National Animal Genome Multistate project were to 1. Develop new genomic tools and assays for more efficient and precise analysis of samples, and 2. Identify and collect liver and blood samples from cows with a history of ketosis and fatty liver and healthy herdmates to provide a unique data set of two phenotypically different populations. The outputs of the first objective include conducting dairy cattle experiments to allow for the collection of liver and blood samples from dry and lactating cattle and use of sequencing technology to sequence and identify SNP variation within target genes. The development of a TaqMan Real-Time PCR Protein Assay was proposed. Thus far, an antibody has been validated and traditional Western Blot analysis has been conducted to allow for validation of the PCR based protein assay. Additionally, we have optimized used of a new Affymetrix GeneAtlas system just acquired by the University of Connecticut Center for Genetics Technologies (CAGT). We will begin using it for bovine whole-genome microarray analysis and will be holding trainings to teach other laboratory groups how to utilize the equipment. The outputs of the second objective include execution of a dairy cattle experiment. The experiment (samples also used under objective 1) included collection of liver and blood samples from dry and lactating cattle, healthy or diagnosed ketosis, from the University of Connecticut dairy herd. These samples were used for sequencing, protein analysis, and liver lipid quantification (n = 16). Several students were involved in these objectives and were mentored by the PI, including five undergraduate students, and two masters students (one non-thesis, one thesis). These students were taught sample collection and processing, techniques, and data analysis. The PI and one of the undergraduate students attended the Joint Annual meetings (ASAS, ADSA) in July 2012 to present research findings. PARTICIPANTS: PI: Heather White Graduate Student, MS level: Sambhu Muraleedharan Pillai, Fall 2012 Graduate Student, non-thesis MS level (no stipend funding provided by this project): Christine McCann Undergraduate Students (no stipend or salary funding provided by this project): Molly Viner, Lisa Dauten, Paulo Zocco, Emma Price, Tia Slivinsky TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Initial outcomes of this project have included learning of new techniques and skills by both the trainees and the PI, fundamental knowledge gained regarding target genes and SNP presence, and adoption of new techniques. The trainees on these projects (both undergraduate and graduate students) have mastered several new techniques. Students are able to collect blood samples and process both serum and plasma and run subsequent assays, surgical preparation for liver biopsy sampling and process liver tissue, isolate RNA, generate cDNA, clone and sequence target regions. The PI and non-thesis masters student have optimized use of the new Affymetrix GeneAtlas system available within the CAGT, and have adopted this as the preferred system for microarray analysis. Additional outcomes have been in the area of specific fundamental knowledge gained regarding a novel target gene, bovine adiponutrin (see publication list). Expression of mRNA and protein abundance reveal that this gene is regulated during the transition to lactation period in dairy cattle. Sequencing and SNP identification of this gene have been performed and potential correlations between SNP presence and metabolism are being examined currently. Further fundamental knowledge is being gained regarding differences in global SNP presence between ketotic and healthy dairy cattle. This data is currently being analyzed.
Publications
- Viner, M. E., S. S. Donkin, and H. M. White. 2012. Hepatic patatin-like phospholipase domain-containing 3 mRNA expression is increased during feed restriction and in transition dairy cows. J. Dairy Sci. 95 (Suppl 2):77.
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