S-STEM Scholar Jessica Sabel's Blog

Introduction:    The goal for this week was to figure out the best ways to grow each bacteria. During the last few weeks, we have had difficulties with multiple species of  Deinococcus . Each species would grow in TGY broth, however, the OD600 values were too low for the species to be used in any testing. This leads us to trying different growth methods to see if there are better ways to obtain a higher yield.    Methods:   400 ul of TGY was prepared and autoclaved. 200 ul of TGY was prepared, separated into 4 125 ul flasks, and then autoclaved. TGY with agar was heated over a hot plate and poured over plates, which were left out overnight to solidify. Media was   Gram staining was done on  D. indicus  and  D. grandis . OD600 values were taken on the nanodrop on  D. roseus ,  D. caeni ,  D. aquatilis ,  D. gobiensis , and  D. indicus . An inoculation was also performed with  D.  caeni onto a TGY agar pl...

Introduction:    Because our  D. grandis  results from 2 weeks ago showed that this bacterium was not resistant at antibiotic concentrations between 1.56 ug/ml and 100 ug/ml for all 3 antibiotics, we are shifting our focus to trying to classify the MIC for the 3 antibiotics we are testing on with an additional 3 antibiotics we have not worked on yet. The goal is to see if  D. grandis  has a higher MIC for all the antibiotics on the board in comparison to the other  Deinococcus  species.   Methods:   400 ml of TGY was prepared following the 3:3:1 method and was autoclaved overnight.   Gram staining was performed on 3/8 on select species shown in figure 3, and OD values were taken on 6 different species of  Deinococcus  on 3/10.   18 test tubes were filled with TGY and autoclaved for 30 minutes. The next day, TGY was removed from each tube to equate to the amount listed next to each test tube in red and purple ink in fi...

  Introduction:  The goal for this week is to complete our protocol for the MIC testing with  D. aquaticus  and start a new procedure with  D. roseus  to identify if it will respond to methicillin, kanamycin, or ofloxacin. We are repeating the same procedures we have done in the previous weeks with both of our tests performed. The main outcome we are hoping for is to identify the MIC for  D. aquaticus,  and to see what antibiotics work with  D. roseus  to see if there are similarities with the previous  Deinococcus  species that underwent antibiotic testing:  D. aquaticus  and  D. grandis .   Methods:    MIC testing with D. aquaticus: 8 test tubes containing TGY with 0.5% agar were autoclaved and set out to cool. The 8 test tubes consisted of: 1 negative control tube, 1 positive control tube with no antibiotic added, 2 3000 ng/ml tubes, 2 2000 ng/ul tubes, and 2 1000 ng/ul tubes. The 2 mg/ml stock ...

Introduction:   Antibiotic testing with  D. grandis  is the main goal for this week. This will be helpful moving forward because it will give us an idea if any  Deinococcus  species respond to these 3 types of antibiotics, and at what concentration it will inhibit growth at.     Methods:   Plates were made from our samples of 25 ml of 1.5% TGY that were in 10 different 125 ml flasks. The flasks were individually heated over a hot plate and removed when all the media was liquified. After the samples were cooled enough to pass the baby bottle test, TGY was removed from each flask so that the final volume in each flask after the tetracycline is added is 25 ml. Tetracycline was then added into each flask and it was swirled to mix. Exact amounts of TGY removed and antibiotic added to each flask  are shown in figure 1. Then the flasks were each poured over a plate and left out overnight so the media could fully solidify. 2 of these samples wer...