On Monday we tested our cultures for carbon utilization. First, a basal media was prepared based on the Luedemann paper, which included 25g/L of yeast abstract, 1g/L of Calcium Carbonate, and 10g/L of NaCl. The 1% NaCl was added to the media because some the stains grew best on marine agar, and also, all the strains had at least a 1% salt tolerance. Control plates were made with just the basal media. Then to the basal media, 16 different carbons sources were added at 1g/L, which included: arabinos (AR),dulictol (DL), galactose (GA), glucose (GU), glycerol (GY), inositol (IN), lactose (LA), mannitol (MN), melezitose (MZ), melibiose (MB), raffinose (RF), rhannose (RH), ribose (RI), sucrose (SU), xylose (XY), and xylan (XN). 10µl of each strain in liquid media was spotted onto the seventeen different plates. The liquid media containing the cultures was prepared by scrapping the cultures from the stalk plates, then washing them with saline to remove any remnants of the previous media including the carbon sources. The growth of the culture on the control will be compared to the different carbon sources. Next, plates were streaked from the stalk plates of the stains that grew at 10ºC to test for growth at 4ºC.
On Wednesday, we first recorded the results from the pH tests. We had many interesting finds. Many of the strains were more tolerant to higher basic pHs than acidic pHs. Also, a few of our strains actually changed color with changing pH. Then we extracted DNA from the dirt samples we used previously in the class. The MoBio soil DNA extraction kit was used with 10g of soil, and the protocol was followed. These DNA samples will be used in a PCR with GEO primers to see if our dirt samples contain any of the studied genera.
The pictures of our pH plates (see those in post of Yellow Rive below) show how stain #69 actually changes color with increasing pH.
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geo primers were used to see if the samples contained species from which genera? im thinking i hear Geodermatophilus but Im not too sure, so enlighten me! thanks
ReplyDeleteOO and im happy someone else forgot, I mean had trouble uploading pictures!!
nevermind i just found my answer!
ReplyDeleteYeah, the Geo primers stands for the family Geodermatophilaceae.
ReplyDeleteIts 2.5g of yeast extract per liter not 25
ReplyDeleteI feel like whenever there is a color change it has been at pH 7 and above and that less color change is seen in the lower pHs. Im not sure about strain 69 but that seems to be a possible trend; however, whether thats significant or even correct or not i dunno, just a thought :)
ReplyDeletewhat evolutionary advantage does expressing different pigment at different pH offer the organism? Could it be that the pigment is always expressed but at different pH's the enzymes that are responsible for their expression are inhibited at different phs?
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