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Notebook Grading

The TAs will look for these items in your notebook. Please complete a self-check before 1pm on Thursday December 8th.

ItemDate in your notebook
Soil collection, GPS recorded 
Initial plaque morphology notes 
2-3 rounds of purification, with notes on plaque morphology and process 
Webbed plate protocol & image 
High titer lysate collection & titer calculation 
DNA purification protocol 
DNA concentration and agarose gel results***AJ will post most recent gels Dec 6th afternoon...sorry it's on my office computer
TEM staining protocol 
Host range testing results- class experiment 
Host range testing results- second experiment 
Y2H mating on non-selective media, pic & your observations 
Y2H selection for diploids on -YT, pic & your observations 
Y2H selection for protein-protein interactions on -YTH, pic & your observations 
Post your contribution to Y2H or host range posters 
Post your final presentation slides 
Post a link to your bacillus.phagesdb.org phage archiving page!! Congratulations, you made it! 

 

 


Important announcement:



Restriction digestion table- please copy to your own wiki page and enter the appropriate volumes.

Solution:1234
10X reaction buffer

2μl

buffer 3

2μl

buffer 4

2μl

EcoRI buffer

2μl

buffer 2

500 ng phage genomic DNA    
10X BSA2μl2μl2μl2μl
ClaI----1μl---------
EcoRI--------2μl----
HindIII------------2μl
H20    
Total20μl20μl20μl20μl

Pipette in this order: Water, 10X buffer, BSA, DNA, enzyme



From 9/8/16 class: 7 successful enrichment cultures! Hope you can see your name and spots.

 


From 9/1/16 class: 19 Successful Enrichment cultures! See if you can see your plates, and the spots on them. A few are really difficult to see, but are there!





Summary Techniques and Data Table: Please copy this table onto your own wiki page. It is your responsibility to notify me or your TA of updates to this table!

Skill
Data
Sign Off


Demonstrate competent pipetting skills

 

 
Demonstrate appropriate aseptic technique

 

 
Phage name  
Soil Sample Collection

GPS:

Physical Description:

Air Temperature:

Collection Date:

 

Discovery method

choose: Direct or Enrichment plating

Enrichment plating

 
Identification of plaque

Date:

Appearance:

 

Purification of plaque

(Number and method used)

 

 

Describe final plaque morphology

(Clear/turbid/halo/comet/size/multiple, etc.)

 

 

 
Production of web plateEmpirical/ Intuitive 
Collection of high-titer lysate

Titer:

Date:

 
DNA purification, how many columns did you use?  
DNA quantification, Concentration/volume:

Concetrantration:

Estimated volume collected:

How is your tube labeled?

 
Restriction digest, which enzymes cut?  
EM Visualization

Head Diameter:

Tail Length:

 

Submitted Archive Report to bacillus.phagesdb.org or

phagesdb.org (Gordonia phages)

 

Insert link here! 

Submitted lysate archive to Dr. Johnson

How is your tube labeled?

 

 

 

 

Our host bacteria

Gordonia terra

Instructors guide for media preparation, etc., for Gordonia

Bacillus thuringiensis kurstaki

 

Samples from Iceland

LocationGPSConditions
Hverir

65°38'25.7"N 16°48'40.1"W

6/30/16, Soil from under edge of one lonely plant in a tuft of grass, at the edge of a sulfur vent area, rainy and 50oF
Glacier Lagoon

64°03'00.7"N 16°10'46.5"W

6/28/16, Grassy area at top of hill overlooking lagoon, sunny and 55oF
Waterfall in east Iceland  

 

Samples from Poland

TownGPSConditions
Poznan52.458737, 16.9105978/2/16, Grassy area beside apartment complex, sunny and mild, 25oC
Bielsko52.608663, 15.927471

8/4/16, Backyard garden of a house, cloudy, 20oC

 

8/9/16 Enrichment cultures

Set up five enrichment cultures containing ~50ml TSB, 0.5 ml Btk overnight culture, a few grams of soil.

Grew at 30oC, overnight.

8/10/16 Spot test enrichment cultures

  1. Prepare plate with lawn of Btk- add 4.5ml top agar to 0.5 ml Btk overnight culture, and pour onto luria agar plate.
  2. Filter enrichment culture sample using 0.22 um filter, into sterile tube. Serial dilute filtrate to 10-5. Spot 3ul of each onto prepared lawn.
  3. Grow overnight at 30oC.

 

 

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