Presentation on theme: "The Effects of PUGNAc on O-GlcNAC Levels and Dauer Formation in C"— Presentation transcript:
1 The Effects of PUGNAc on O-GlcNAC Levels and Dauer Formation in C The Effects of PUGNAc on O-GlcNAC Levels and Dauer Formation in C. elegansBy Nicole Kiess
2 NeedPicture 1: Shows the rate of Type 2 Diabetes in relationship to obesity over 10 years.
3 NeedEstimated prevalence of diagnosed and undiagnosed diabetes in people aged 20 years or older, by age group, United States, 2007Picture 2: Shows percent of people in different age groups with Type 2 Diabetes.
4 Knowledge BasePicture 3: Is a model of what happends in the body of a Type 2 Diabetic
5 Knowledge BasePicture 4a: Shows the pathway that regulate aging in C. Elegans.Picture4 b: Shows the model for the regulation of life span and sensory perception.
6 Knowledge Base Picture 5: (Top) Insulin-like signaling pathway. (Bottom) C. Elegnas life cycle.
7 Literature ReviewParadis, et al,(1998) showed that mg144 does not activate AGE-1 P13K and that Akt/PBK is not dependent on AGE-1/P13KPicture 6a: Shows normal growth conditionsPicture 6b: Shows dauer formation conditions
8 Literature ReviewArias, et al, 2004 showed that PUGNAc increases O-GlcNAc levels in rat skeletal muscle
9 Literature ReviewHanover, et al, 2005, findings show that OGT modulates macronutrient storage and dauer formation in C. ElegansPicture 7a: Insulin like signaling pathwayPicture 7b: Shows percentages of dauers in different strains
10 Literature ReviewForsythe, et al, 2006 demonstrated that enzymes of O-GlcNAc cycling perfect insulin-like signaling in response to nutrient fluxPicture 8a: Insulin like signaling pathwayPicture 8b: Dauer formation in different strains with temperature as a variable.
11 PurposeTherefore the purpose of this experiment is to observe the O-GlcNAc levels, using densinometry, and dauer formations with PUGNAc as a variable.
12 HypothesisNull- There will be no change in O-GlcNAc levels and no change in dauer formationAlternative- The levels of O-GlcNAc will rise and dauer formation will rise
13 MethodThe effects of PUGNAc on O-GlcNAC levels and dauer fomation in the insulin-like signaling pathway of C. elegansControlWild Type N2Ogt-1 (ok430)Oga-1 (ok1207)Daf-2 (e1370)50um100um50um100um50um100um50um100umC. Elegans will be cultured at 16 degrees CO-GlcNAC levels will be observed by treating C. elegans with RL1 and RL2 antibodiesPictures will be taken with camera under immunofluorescent microscope and photoshop will measure percentagesAdults of each strain lay embryos overnight at 15 degrees C. Cultures shifted to 22.5 degrees C and are scored 3 days later as either dauer or non dauerStatistical Analysis will be run with SPSS and an ANOVA
14 ProtocolC. elegans will be cultured in petri dishes with the Nematode Growth Medium (NGM) and will be fed Escherichia coli (E. coli)Picture 9: How the C. elegans will be culturedDrawn by Victoria Wei
15 Protocol RL1 and RL2 dilated 1:400 and given to C. Elegans Picture 10: Life cycle of C. elegans
16 Protocol Fluorescent microscope and camera used for picture Photoshop used for percentagesPicture 11: A) Fluorescent picture to show O-GlcNAc levels with RL-2B) Fluorescent picture to show O-GlcNAc levels with RL-2Hanover, et al, 2005
17 Do-Ability Need to Purchase: Already Obtained RL-1 RL-2 C. elegans strainsE. coliPUGNAcAlready ObtainedFluorescent microscopeCameraPhotoshopPetri Dishes
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