BioTech Lab Entry 4

 Introduction

    This week we performed performed our SIM test and the UV sensitivity test on Deinococcus xinjiangensis. 


Methods

    On Monday, SIM media is inoculated with E. coli, Staphylococcus epidermis, and D. xinjiangensis by use of an inoculation needle. Two gram-stains are performed of D. xinjiangensisi, one of which is negative and the other is inconclusive. Two TGY plates are inoculated with D. xinjiangensis. The plates for the citrate test are checked for the presence of blue. Old SIM media from the previous week is discarded. On the following day, two gram-stains are performed on D. xinjiangensis grown on TGY. The results of thests stains are both gram-positive. Two 300mL flasks of R2A are prepared and used to pour 20 plates, two of which are inoculated with D. xinjiangensis. On Wednesday two gram stains are done for the R2A plates grown yesterday, which are both gram positive. The following day OD values are taken for two flasks of R2B and one flask of TGY of D. xinjiangensis. 1000uL of these flasks are then put into three separate epindorph tubes. 10 plates were labeled with sharpie and lines were drawn which separated the plate into three separate components. 50uL of bacteria were exposed to increasing intervals of 1000 units of UV radiation, starting at 1000 units and ending at 9999 units (10,000 units). The bacteria were then streaked onto plates of R2A and incubated at 30 degrees Celsius. This procedure is repeated on Friday but for bacteria cultures grown in TGY instead of R2A. However, due to issues with dilution, we were unable to perform the UV test.


Results

     Deinococcus xinjiangensis tested negative for the ability to break down citrate. The results of the UV test will not be obtained until the following week.


Discussion

    It is reassuring that the gram stains came back positive. Potential sources of error for the previous gram stains were leaving the safranin on the plate for too long, giving too long of an ethanol wash, or possibly bacteria grown on R2A are compromised somehow. Further experimentation is needed to determine the cause of the discrpency. 

Comments

Post a Comment

Popular posts from this blog

BioTech Lab Entry 6

  Introduction     This week was spent desiging ways to further test the UV resistance of Deinococcus xinjiangensis .  Methods    Six flasks of TGY, each containing 25uL, are prepared. Five flasks of 20 uL of R2B and one flask of 20 uL of R2A is prepared. A practice gel is done for electrophoresis. The gel contains 30mL of TAE buffer and 0.3g of agarose. Cryovials contain 2uL of the sample, 8uL of PCR water, and 2uL of the loading dye. The gel is run at 100 amps. The next day, 2 flasks of TGY and one flask of R2B are inoculated with D. xinjinangensis . They are put in the skaing incubator at 30.0 degrees Celsius. I then had a discussion with Dr. Tuohy on further avenues of research (see discussion). Gram stains are done on the flasks inoculated yesterday. However, the results are inconclusive. Aland and I design a new and simplified experimental design for the UV tests that can be easily manipulated to test for specific variables. The following day, A...
Read more

BioTech Lab Entry 7

  Introduction     This week, a UV test was performed on Deinococcus xinjiangensis . Competent Deinococcus sonorensis  were made and a plasmid extract was done on D. sonorensis  too.  Methods     On Monday, Gram stains are performed on D. xinjiangensis  grown on three different plates: one of R2A, and two of TGY. The bacteria grown on TGY are gram-positive, while the stain done from R2A reveals contamination by Staphylococcus . Two TGY plates and two R2A plates are inoculated with D. xinjiangensis . 2 flasks of 150mL of R2A are also inoculated with D. xinjiangensis . Two R2A plates are inoculated with D. caeni . The following day, the plates inoculated with D. xinjiangensis  are tested through a gram-stain. All of the results are gram-positive. A 200mL flask of R2A is made and OD values are taken of the D. zonorensis  inoculated yesterday. On 4/24/24, three 2mL centrifuge tubes containing R2B and D. sonorensis  are centrifuged ...
Read more

GCC BioTech Lab Blog Entry 2

Introduction     This week in the lab was not very busy. We hoped to innoculate plates and perform characterization tests on Deinococcus xinjiangensis , but were unable to do so. Our group also began investigating whether or not certain proteins involved in mechanotaxis and chemotaxis were present in various Deinococcus  species.  Methods       28 plates were prepared. 10 contained TGY, two contained a comibination of starch and TGY, six contained urease, and 10 contained R2A. 500 ml of TGy was heated at 350 C and poured into the TGY plates. Another TGY flask of the same volume was heated at 355 C and poured. 2g/L of strach and TGY was used to make the starch plates, while the solution for the urease test consisted of 50mL of urease in 450mL of H20 and Agar after the latter solution was heated to 350 C. After the plates were poured they were placed in the refrigerator. Next a R2B broth was prepared. 0.378g of R2B powder was dissolved in 120 mL of DI wa...
Read more