1、Study Plan Study Plan mainly includes:1. Study background: the current research condition and level of the study subject at home and abroad2. The purpose of studying abroad, the anticipated objective, the plan, the method of implementation and the time needed3. The feasibility of attaining the antic
2、ipated objective (combining it with the professional technology and knowledge that you have mastered ) 4. The foreign country and the academic institution that you have chosen, and the reasons (please simply narrate the advantages and level of the academic institution abroad, and if there are cooper
3、ation foundations and contacts between your present institution and the institution abroad. If there is an offer, please add a copy ) 5. The study (working) plan in China after finishing the study at abroadStudy PlanPersonal status: I am *, female, born on 24th of Dec., 19*. Ive been dreaming to a b
4、ig figure in the circle of business. So when I graduated from senior high school, I chose to pursue professional knowledge about business. In September 2002, I was enrolled to study in * School in Beijing, with major of Business Administration (in English) And then in March 2005, I successfully prog
5、ressed into * in Beijing, the associated university*. I have mainly studied the following subjects: English, Mathematics, Computer, Business Law, and Principle of Accounting, Finance and so on.The Reason for My Choice of Studying in the Canada: I have a small but warm family. My parents are very suc
6、cessful in their careers. My father is * and my mother is *. I am proud of them. I regard them as my role model. I make my resolution to study hard for standing out in the future.After three years of study, I realize that it is more challenging than I had imagined, which makes me more attracted to i
7、t. But I found myself lack of advanced knowledge business. So to study abroad emerges in my minds. Luckily, my parents are very supportive and are willing to financially sponsor my study in the Canada. Through comparison, I finally choose to study in the Canada because of her famous education, beaut
8、iful scene and safe living place.The Reason for Choosing *(你要去的学校的介绍):The Proposed Courses: I have been accepted to study * Programme in * as follows:Course: * Degree programmeTotal credits per year: 30 creditsStart Date: *Completion Date: *After obtaining bachelor degree from Canada, I plan to come
9、 back to China without hesitation to put all the principles I have got into practice. China has witnessed a rapid growth in economics, so Ill have lots of opportunity to develop myself based on my golden diploma and proficient English. Accumulated rich experiece, I really want to have my own enterpr
10、ise in order to prove my ability, which are also my parents willing. The more importanc is that I love my parents. I am the only child. I think I should reward them with my outstanding achievement. And Id like to share every step of success with my dear parents.Respectful officer, if I am luck to ge
11、t the permission to go to Canada to further my study, which I believe is the first step to approach my goal, I will treasure this opportunity, make every effort to improve myself. Thank you.Signature of the Applicant:Date of Signature:A sample study planStructural and Functional Studies of Human Sun
12、1 OBJECTIVES To determine the orientation of Sun 1 in biological membranes. To express human Sun 1 in E. coli. To purify and refold the recombinant protein . To study human Sun 1 s 3D structure and molecular interactions by NMR or X-ray crystallography. OVERVIEW OF THE STUDY Human Sun 1 s 3D structu
13、re and its interaction with other components in the nuclei are unknown. Recombinant proteins are popular choices in studying such issues. We plan to express human Sun 1 in a suitable Escherichia coli expression system which has several advantages over other expression systems, such as no post-transl
14、ational modifications like glycosylation. By different approaches, such as mutant selection, protein fusion, using protease-deficient strains, etc., we can overcome toxicity and proteolysis which are often associated with expression of membrane proteins . If human Sun1 is expressed directly in the E
15、. coli membrane, we could obtain valuable membrane models to study Sun 1 s orientation and in vivo interactions with other molecules. Next we will purify the recombinant protein by affinity chromatography which is one of the two suitable methods to study membrane proteins. After the purification, we
16、 will refold the protein with the aid of micelles or liposomes in order to prepare samples for NMR study. S tudies have shown that Sun1 is detergent, salt, and chaotrope resistant . Fortunately, it is also shown that treatment with detergent and medium salt concentrations (1% Triton X-100/ 200 mM Na
17、Cl) can efficiently solubilize Sun1, which can be a good start for our study. Currently, membrane proteins can be studied either by NMR or X-ray crystallography. However, for NMR, only protein fragments with molecular weight less than 20 kDa can be studied. For the limitation of NMR, we can study in
18、dependently the transmembrane domains of Sun1 by NMR, and combine the experimental data, which can sometimes give sufficient information about the target protein. EXPERIMENTAL PROCEDURE Sun1 polyclonal antibodies Synthesize a peptide having the first 14 amino acid residues of human Sun1 (MDFSRLHMYSP
19、PQC; NP_079430 ). Sun1 polyclonal antibodies will be produced against this peptide in rabbit. The anti-Sun1 serum IgG fraction (referred as the Sun1 serum) will be purified on a protein A-Sepharose column. 2. Recombinant Proteins Sun1 was initially isolated from human brain. SUN will be PCR-amplifie
20、d from the Sun1 cDNA and directly cloned in pcDNA3.1 TOPO/V5-His and pET21 vectors. Recombinant pET vectors will be transformed into BL21(DE3)plysS host which is ideally stringent to express membrane proteins. Verify if recombinant protein is expressed in inclusion bodies and purify it by Ni column.
21、 3. Gel electrophoresis and Immunoblotting Test the dilution first and then use the Sun1 serum as the primary antibody. 4. Proteinase K Protection Assays The Sun1 cDNA cloned in the pcDNA3.1/V5-His vector will be used to synthesize the full-length protein in vitro with 35 Smethionine in the absence
22、or the presence of canine-pancreatic microsomal membranes. Aliquots of the synthesis reaction will be digested for 30 min on ice with proteinase K. Proteinase K will be inhibited with PMSF, and the samples will be added directly to boiling Laemmli buffer. Duplicate samples will be transferred to a n
23、itrocellulose membrane, and the radiolabeled proteins will be visualized with a PhosphorImager. The same membranes then will be immunoblotted with either the anti-Sun1 serum or the anti-V5 antibody. 5. Reconstitute Recombinant Protein Fragment Functionally into Detergent Micelles In order to prepare
24、 samples for NMR study, we will first choose suitable detergent micelles to reconstitute the recombinant protein fragment , use a direct dilution method to refold the proteins, and test the refolding of the target protein by radio-ligand binding. 6. NMR and X-ray Crystallography Studies We will exam
25、ine whether solution NMR studies of the micelle systems ( using different domains of the target protein ) will give sufficient information about the structures of human Sun1. As for X-ray crystallography studies of the whole protein , we will choose a suitable detergent first to work out protein cry
26、stals and then perform structural studies on the crystals . FURTHER STUDIES With the aid of NMR and X-ray crystallography, we can study directly protein-drug interactions and protein-protein interactions. We will predict human Sun 1 s 3D structure by molecular modeling and compare it with experimental data from NMR or X-ray crystallography.
