Mentors+Fall+13

MENTORS: DANIEL, MICHAEL, ANDREW, URVASHI, MAX, JANICE, BRANDON, SUMAN, ALDO, DIVYA

Please Enter your class schedules on the Google Docs Spreadsheet
 * Recurring Tasks:**

Mentors, use the MentorLabLog to record research related tasks that you have done so that we can have continuity when passing off a task from one mentor to another e.g. transformations, bacterial cultures, conditions of protein preps, storage conditions, etc.

Find some good journals and link their RSS feeds to a new page on this wikispaces called Current Literature

Tasks for 15th Week Fall 2013 (Dec2nd - 7th)
Total Lab hours and enter onto spreadsheet - done Andrewski

upload to Docs/VDSstaff/Grading_Wiki_LabNotebook_JournalClub_Etc/M&Ms_PeerReviewChecks At the top row of the spreadsheet are the Due Dates for each assignment
 * M&M's** **Check**
 * **M&M Cloning - DONE**
 * **M&M Virtual - DONE**
 * **M&M Expression, Purification, Characterization - DONE**
 * -** determine who has turned them in late (or not at all)

Log In to Canvas course go to the Grades tab for the each M&M -- if the box is pink - the assignment was late, click on it to see when the submitted, then enter the # of days late onto the spreadsheet. If they have an excused late - subtract those dates from the total days late. -- if the box is missing, then type 'missing' and highlight the cell red, enter '100' for # of days late


 * Peer Reviews check**
 * ** Peer Review for M&M Cloning - ** done by Sumanski, Urvashiski, and Maximillianoski
 * ** Peer Review for M&M Virtual **
 * ** Peer Review for M&M Expression, Purification, Characterization **

- login to Canvas Click on 'Assignments' then on the M&M that you want to take you to the setup page. - on the right, click on 'Peer Reviews' Keep this page open on one side of your screen

Then open the other side of your screen with the 'Grades', then for the assignment you want, mouse over the blue name of the assignment until you see a little down arrow - to open 'Speed Grader' -- verify that the Peer Reviewer has made comments for both of their papers. -- Just becasue they have a yellow '!' mark on the Peer Review page, doesn't mean that they haven't done it. Sometime they put them in the 'Discussion' section, sometimes they upload a comments file, and sometimes they are just on the yellow comment boxes on the page. -- if they did not do one of their reviews type: '0' and make the box red -- if they did not make enough comments (5 criticisms and 2 compliments) then: give them an appropriate score out of 10. -- if they could not do their because the other person did not submit - then only grade the one review. But make a note of it somewhere.

Pfu Polymerase tests - Michael.

Grade Journal Club - Study Q's & Journal Club Group Q's - after the Wednesday class

Tasks for 16th Week MONDAY only (Dec9th))
Total Lab hours and enter onto spreadsheet - Check Wikispaces (use Google Docs Spreadsheet that we have) - Research Progress spreadsheet updating. In the /VDSStaff/LabHours folder on GDocs -- LabProgress -- use Wikispaces and LabNotebooks to determine their progress --

Verify that students have done their 2 Peer Reviews of other students' M&M Cloning. - -This is on Canvas. -Create a new Column in the Wikispaces grading sheet -Check to see if they have given: --5 Constructive Criticisms --2 Compliments Give 10 pts for a good review, give 9.5 for an ok review, give 9 for a not good review, give 0 for nothing.

TASKS FOR 14th WEEK (Turkey Week):
dilute the phosphatase compounds - to 50 mM in DMSO - to 5 mM in DMSO/Water See the instructions diluting compounds in GDocs/Misc/Compounds

Make some 0.01 mM Orthovanadate in water

Make some tubes of 50% water/50% DMSO. Make 20 tubes of 10 ul each in the 0.600 ml tube size ( medium sized micro centrifuge tubes)

TASKS FOR 13th WEEK (Nov 18th - Nov 23rd):
Total Lab hours and enter onto spreadsheet - DONE Dilute Compounds - (see Below) - started by Suman Transform YopH in pNIC-Bsa4 into BL21(DE3) cells - done by Urvashi and Suman -- NEED to check plates for these and Transfer to 4degC - TUESDAY Mentors do this......

Make 20% Sucrose - sterile filtered - Suman ** -- DONE ** Degass some FPLC buffer - Suman ** -- DONE **

Make LB Agar/Kan/Suc plates - TUESDAY Mentors

Check Wikispaces (use Google Docs Spreadsheet that we have) - TUESDAY Mentors

Michael made the buffer calculations for Pfu polymerase - see paper sheet.

TASKS FOR 12th WEEK (Nov 11th - Nov 16th):
Total Lab hours and enter onto spreadsheet on GDocs, put Lab Hours sheet in Dr. B's 'reserved' cubby when done. **-- DONE**

Research Progress spreadsheet updating. In the /VDSStaff/LabHours folder on GDocs -- LabProgressAsOf111113 **- DONE** -- use Wikispaces and LabNotebooks to determine their progress --**DONE**

-This is on Canvas. -Create a new Column in the Wikispaces grading sheet -Check to see if they have given: --5 Constructive Criticisms --2 Compliments Give 10 pts for a good review, give 9.5 for an ok review, give 9 for a not good review, give 0 for nothing.
 * Verify that students have done their 2 Peer Reviews of other students' M&M Cloning. -**

go with the new students when they are doing sonication and big centrifuge. You should go with them unless they have done it 2 times on their own.
 * Sonicator & Big Centrifuge:**

Also, please go in and check on the big centrifuge in the Brown lab (Bioprospecting) next door to make sure it hasn't been left on or the door has been left closed after it is done. -- we have had a problem with the fridge being left open.

Make a picture protocol -- for the Autoclave - go take pictures of open and closed door of right autoclave, pics of left autoclave, pics of screen on left autoclave

-- for the new big centrifuge. Take pics of open and closed and of the screen with the correct '10' or '20' entered for rotor type. Take pics of fridge door.

Run them by Dr. B for revisions Post both protocols to our GDocs, but also post the Big Centrifuge one on the actual centrifuge.

-- 500 ml of TGS 5X stock buffer for running gels. See the Spring 2013 Buffers&Solutions protocol in the **GDocs/Protocols/Sp13 Labs -- DONE**
 * Make TGS**

-- make 2 Liters of LB and keep in bottles in 4degC fridge. Label as "VDS Class" and put a date on it. ** -- DONE **
 * Make LB**

Use this **Google Docs/Misc/Compounds/instructionsdilutingcompounds** for instructions & sheet to do calculations -- STEP A: Make 50mM Stocks of each compound in 100% DMSO and enter info on the table -- STEP B: do dilution test of compound in a mixture of DMSO/Water -- STEP C: Make a final dilution for the students' enzymes assays using the concentration that worked - i.e. was soluble
 * Dilute Compounds when they arrive from Chembridge:**

TASKS FOR 11th WEEK (Nov 4th - Nov 9th):
PyMol – Schrodinger website has the citation, Cite the original GOLD paper – the 1995 one– see the CCDC website SWISS-MODEL, ICM Molsoft, MolProbity - **DONE**
 * Wednesday 11/06/13**
 * Need a mentor to find the webpages that have the 'proper' journal paper citations for these resources - i.e. not just a webpage citation. Then list them in Piazza as a post under M&M lablel.**

Wikispaces checks - complete theses

Lab Notebook checks - (veterans book only)

Test Pfu Polymerase using PCR for pNIC-Bsa4 protocol. -- use Q5 buffer, pLic For and pLic Rev primers

TASKS FOR 10th WEEK (Oct 28th - Nov 2nd):
make overnight cultures (4 colonies - of FtHAP) in 4 ml of LB - pass of the overnight culture to next mentor also grow up overnight cultures of Pfu polymerase cells
 * Tues**

Make Glycerol stocks of cultures and store in -80degC (metal basket). Label well. - see protocol on GDocs for glycerol stocks
 * Wed**

keep notes in MentorLabLog - to record what has been done AMP plate in Fridge (we think they are BL21(DE3) cells grow in up in 5ml LB and make glycerol stocks for archival storage Grow 2 liters in BL21(DE3) and express, purify, characterize - see protocol in VDSStaff GDocs
 * grow up Pfu polymerase (obtained from Dr. McDonald), -** Daniel, Michael, Andrew

Test batch of Pfu polymerase - PCR amplify a known target (e.g. pGBR22), or the Sac B gene.

TASKS FOR 9th WEEK (Oct 21st - Oct 26th):
Thursday - check the overnight cultures (6 colonies - from 3 plates - FtHAP, YopH) Plates labelled A,B,C and colonies labelled A,B,C

if they grew up make 'ok' and the date on the plates. If they did not grow up - discard the plate (but don't throw away Joey's FtHap - just write on it 'bad" and the date)

Maybe: Make Glycerol stocks of each if they grew up and store in -80degC.

Comments on the Mid-Semester Research Reports assigned to you (~6-7) - in PROGRESS -- use Canvas 'Sandbox' site to do Speed Grader. (collaborative reviews)

Lab Notebook grading (Springers) - DONE (Dr. B)

In Order of Priority 1. Get in touch with your Target Team for a meeting. 2. Total Hours and enter on spreadsheet GDocs DONE - THANKS 3. Wikispaces checks and grades DONE - THANK YOU 4. LIC cloning Exercise grading DONE- THANK YOU 5. Distribute Journal Club papers DONE- THANKS

create a MentorLabLog on VDSstaff account of GDocs/Misc folder - to record what has been done AMP plate in Fridge (we think they are BL21(DE3) cells grow in up in 5ml LB and make glycerol stocks for archival storage - see protocol on GDocs for glycerol stocks Grow 2 liters in BL21(DE3) and express, purify, characterize - see protocol in VDSStaff GDocs
 * grow up Pfu polymerase (obtained from Dr. McDonald), -** Daniel, Michael, Andrew

Test batch of Pfu polymerase - PCR amplify a known target (e.g. pGBR22), or the Sac B gene.

TASKS FOR 7th WEEK (Oct 7th - Oct 12th):
-- total lab hours from paper sign in sheets and upload to spreadsheet -- verify lab safety training for all students --create Target Groups + Mentor in Piazza, send announcement email prepare FPLC - buffer. Make 50 mM Tris, 150 NaCl, pH 8.0 sterile filter, degass see /GDocs/Misc/SolutionsandDilutions
 * Mon** -
 * Tues** -
 * Wed** - add rest of Journal club papers to Mendeley account (vdsclass@gmail.com, you know the password)
 * Thurs**
 * Fri**

NOTE: for the next Journal Club on the Ricin paper from Dr. Robertus' lab, I have placed an example image of what their PyMol answer for Question #0 should roughly look like. See the handout Q's on the Journal Club page for specifics in GDocs/VDSStaff/Misc

TASKS FOR 6th WEEK (Sept 30th - Oct 5th):
-- total lab hours from paper sign in sheets and upload to spreadsheet -- verify lab safety training for all students prepare 5% sucrose, sterile filtered
 * Mon** - prepare 5% sucrose, sterile filtered
 * Tues** - prepare 20 Kan/Suc/Plates - I think we have alot
 * Wed** - journal club grading, add rest of Journal club papers to Mendeley account (vdsclass@gmail.com, you know the password)
 * Thurs**

prepare FPLC - buffer. Make 50 mM Tris, 150 NaCl, pH 8.0 sterile filter, degass see /GDocs/Misc/SolutionsandDilutions


 * Fri**

TASKS FOR 5th WEEK (Sept 23rd - 28th):
Friday - verify Training History pages (paper versions in folder) of Springers. Check of which ones have everything on the Safety page **- Aldo** use class sign in sheet to see list of all students http://vdsstream.wikispaces.com/Required+Lab+Safety

Create a Posters folder in our **Mendeley** account and create citations for our VDS posters - **Brandon & Max**

Dr. B posted the Word formats of these in VDSStaff/GDocs/Misc

Test out: LICpNIC-Bsa4cloningExerciseVDS_Fall13.docx -in VDSStaff GDocs/Misc folder - **ALL Mentors. Daniel, Michael and Andrew did this and summer - so ask them how**

TASKS FOR 4th WEEK (Sept 16th - 21st):
Test out: Virtual Screen Refresher - (not posted yet -0911113 Dr. B) - **Brandon & Urvashi**

TASKS FOR 3rd WEEK (Sept 9 - 14th):
Complete the Weeks Layout on the Research Pages page: **Michael - n/a** http://vdsstream.wikispaces.com/Research+Pages+2013 Continue making pGBR22 - **Brandon & Suman - Suman and Urvashi completed (091013) - Thanks - Dr. B**

Thursday & Friday - Concentrate and store in -20degC w/ 20% glycerol - YopH protein sample (that Janice & Urvashi made) - **Suman & Max - DONE - Thanks! - Dr. B** - 4 vials of 20% glycerol stored in VDSStaff box in -20degC - a few vials of snap frozen stored in -80degC in a cardboard box (smaller size box)

TASKS FOR 2nd WEEK (Sept 2 - 7th):
Friday or Thursday - edit new Hours Column Chart Fall 13 graph and link to Wiki - **Urvashi - get MICHAEL's help**

Friday 09/06/13 - **Urvashi** - total students' hours from the sign in sheet and upload to the **Hours Column Chart Fall 13 (in the VDSStaff account)**

Purify YopH and run on gel **- Urvashi Done Purifying** Gel check YopH samples - **Janice & Urvashi**

Grow up 2x 160ml of **pGBR22** in DH5alpha for the students' RE Digest protocol (check if we have plates first - may need to do transformation), next day spin down and Midi-prep, split into 2 tubes, Nanodrop, send to DNA Sequencing - **SUMAN, BRANDON** NOTE ($): When doing Submit to DNA sequencing - we have a NEW account number. Be sure you don't use the old one from last year. NOTE: newly made plasmids go in the **'Unverified Plasmids'** box until they have been verified by DNA Sequencing.

TASKS FOR 1st WEEK:
Take a new set of pictures of the mentors with their names - to put on the this Mentors page. Take groups of 2 or 3 or use individuals. - **Suman**

schedule mentor meeting - **Urvashi** DONE - thanks! prepare mentor schedule on GDocs - **Suman** DONE - thanks! prepare student schedule on GDocs - **Suman** DONE - thanks! make Mendeley account and upload papers - **Brandon** DONE - thanks! update VDS google Website and upload this past years posters as PDFs - **Michael** DONE

Piazza - **Urvashi** -- add link to Left hand table of Wiki to Piazza class - DONE - thanks! -- add all enrolled students to Piazza. - DONE - thanks! -- create Target groups in Piazza to allow within-group communication (we know the summer groups already on the Targets page) - 2-4 students + Mentor -- also create Veterans and Springers groups DONE

Create new 'blank' pages on the @Research Pages 2013 for each new student. Organize them into Veterans, Springers - **MICHAEL** - DONE - thanks!

Prepare MENTORS folder in lab:

 * 1) print new timesheet pay periods sheet (from UT webpage - see link below) **- DANIEL**
 * 2) create and print new hourly sign-in sheet - **ANDREW**
 * 3) get some extra blue sheets to include in folder - **ANDREW**

Walk through TDRTargets Exercise - on computer - **Max, Suman, Aldo, Janice, Brandon, Urvashi - n/a**

Generate assessment quiz questions over Spring Material - two mentors team up -
 * Daniel & Michael**

Generate assessment quiz over cloning - two mentors team up -
 * Andrew & Urvashi DONE**

Update Laptops and Desktops - ANDREW, MICHAEL - DONE

 * Start up laptops and do updates on them. Remove security credentials. - ANDREW, MICHAEL **DONE**
 * Clean off old icons from Desktops of Desktop computers - ANDREW, MICHAEL **DONE**
 * Do Windows Updates - ANDREW, MICHAEL **DONE**
 * Install both printers on each laptop and each Desktop (see below) - ANDREW, MICHAEL **DONE**

WET LAB WORK
Prep collirollers - split into 2 bottles and rinse then autoclave, then distribute ~6 beads/tube into 600 ul microcentrifuge tubes in a sterile fashion and store in 96well plastic 'brick' in our cubby. - **ALDO** DONE - thanks!

1 person make a 100 bp ladder (need to have received it from ordering) - **ALDO** DONE - thanks! clean pH probe and bottle and replace solution - **ALDO** DONE - thanks! label reagents that were in the -20degC - Q5 polymerase and BsaI - put VDS and date and store in Top-Loading freezer - **ALDO** DONE - thanks!

Make some Kan plates (~50) - **SUMAN, BRANDON** - DONE - thanks!

Make autoclaved water and aliquot several into 1.7 ml centrifuge tube, label and store in -20degC. - **SUMAN and BRANDON**

Transform YopH into BL21(DE3) - **JANICE, MAX** DONE - thanks!

Test out and revise overnight expression protocol using YopH - **JANICE, MAX** IN PROGRESS

general lab cleaning - **URVASHI //done//** put specs in correct boxes, see if we are missing anything - **URVASHI** //(done. note: Rugsby and Thomas are missing power supply)// Get rid of old LB media and wash bottles - **URVASHI** //done// organize red Sigma Kit boxes (PCR Cleanup, Gel Extraction, Miniprep Kit). Consolidate and then discard all but two boxes for each type: current box and a spare solutions box. - **URVASHI //done//** Organize plates by Staff, vs. Summer, vs etc. - **URVASHI** -- Put contaminated plates in zip locs, mark for disposal, but leave in fridge for the user to dispose of (I want them to see first).

VDS Staff Meeting: 1:30 on Tuesday August 27th.

 * On Wikispaces page**

- in the back of the lab (on top of fire cabinet) - use plastic sheet protectors
 * Organize and print new protocols (updated) for the white protocols binder**

PURCHASING at BioScience Storeroom (NHB)
spray bottles - 2

FOR LATER:
send pGBR22 samples to DNA Sequencing so that we can figure out which ones had the contamination (maybe have the students do this)

Clean water baths (the big ones - probably need 2 people for this). Only use Alconox soap - no bleach or anything harsh.

Check lab safety completion for all students
-- see the printed sheets in the lab binder and then cross check them to the Wikispaces 'Required Lab Safety' page - cross reference to UTEIDs of Class Roster (can be found in VDSstaff@gmail.com account

**Check Ice Room access list -**

= Printer Installation =


 * 1) Login as Mentor, Start Menu,
 * 2) Devices And printers
 * 3) Add New Printer
 * 4) Add "local" printer
 * 5) Create New Port --> standard TCP pathway
 * 6) Enter IP 128.82.195.186
 * 7) Printer Name Lexmark C543
 * 8) Add Printer (if can't find printer do Window Update)


 * For installation of a different Printer (such as Dell) change the IP address and Printer Name**

Printer Name: Lexmark C543 IP: 128.83.195.186

Printer Name: Dell IP: 129.116.152.49

**SOLUTION RECIPES**

 * Make LB** - 2Liters - and autoclave


 * Make 5 M NaCl** - 500 ml in water, sterile filter into a bottle, label, store at Room Temp


 * Make Kanamycin**- stocks 50ug/ul store in -20degC, Make 30 tubes of 500 ul each. __Sterile filter with syringe filter__before aliquoting into tubes. Label. Store in -20degC


 * Make IPTG** - 1 M IPTG stock to make. Make 30 tubes of this with 500 ul in each. Label. Store in -20degC


 * Make Lysozyme** - 20 aliquots of 500 ul at 50 mg/ml in 1.7 ml tubes. Label. Store in -20degC


 * Make TCEP -** make ~50 x 100ul aliquots of 500 mM aliquots medium size centrifuge tubes. Label, put in a box and Store at Room Temp in cubby.


 * Make PMSF -** 30 x 500 ul aliquots of 10mM in **__Isop____ropanol__** 1.7 ml tubes. Label. store in -20degC


 * Make Imidazole** - 1M stock **make in beaker of 500 ml of water, then bottle top sterile filter it into a bottle, store at Room Temp**

Lysis Buffer for Sonication (make 500 ml)

 * (see GDocs '**Solutions&Dilutions**' in /**Misc **folder for calculations)**

Tris 100 mM

NaCl 300 mM

Imidazole 30 mM

pH 8.0 w/ NaOH

label, store at 4degC


 * NEW Wash and Elution Buffers and Storage Buffer(see GDocs '**Solutions&Dilutions**' in /**Misc **folder for calculations)**

Wash Buffer

Tris 100 mM

NaCl 300 mM

Imidazole 30 mM

pH 8.0 w/ NaOH


 * Elution Buffer**

Tris 100 mM

NaCl 300 mM

Imidazole 300 mM

pH 8.0 w/ NaOH


 * FPLC column buffer - tris Also 'storage' buffer**

Tris 50 mM

NaCl 150 mM

pH 8.0 w/ NaOH

Then filter sterilize

Then degass**

6X SDS Page Buffer from 5 Prime Protocol
30 ml of glycerol – add first so that you can see the volume (viscous)

(In a separate tube) - 30 ml of Nanopure water

--- 1.635 g Tris powder

--- 3 g SDS poweder

--- 0.3 g Bromophenol blue

2.3136 g DTT

Mix contents of second tube into tube with glycerol

Add some more Nanopure water to 50 ml total final volume