Introduction
Background
This experiment pertained to identifying an unknown bacterium. This bacterium was retrieved from a member of our group’s hand. Bacteria are prokaryotic, but like all other living organism there are many genus and species. Through experimentation these unknowns can be found can be brought to light. Bacteria can be classified either gram-positive or gram-negative. These groups differ by the composition of their cell walls. In particular, the difference in the composition pertains to amount of peptidoglycan, a polymer of amino acids and sugars, present. A Gram test is most commonly used for this classification, and pertains to using a dye to determine the classification of the bacteria. The peptidoglycan that is found …show more content…
A KOH string test is one of these methods. In this method, the bacteria is mixed with KOH, and while mixing the sample will either become viscous somewhat quickly (Positive String) or it will not (Negative String). This reaction correlates with gram-positive or gram-negative classification in a somewhat opposite manner. A positive string test indicates a gram-negative bacteria and a negative string test indicates a gram-positive bacteria. Another method that can classify gram-positive or gram-negative is the presence of bacterial growth on different media. Eosin Methlyene Blue-lactose (EMB-lactose) is an example of a media that allows gram-negative bacteria to grow and …show more content…
PCR is a common practice used to amplify a certain region of DNA. In this experiment, PCR was used to amplify the 16S ribosomal RNA gene of the bacteria. This locus was used because it is one that evolves slowly in bacteria. A Taq polymerase is used for this PCR because of the temperatures the reaction goes through. The16S rRNA primers used in this PCR, are complimentary to sequences conserved in all bacteria and will target sites of variation. These sites of variation were the regions of the DNA amplified by PCR. The products of PCR cannot be sequenced correctly without purification. Purification removes left over reactants and enzymes. A spin column-collection tube complex is used to collect the DNA during a series of centrifugation techniques with different buffers. The silica membrane holds DNA in a high-salt buffer and will release it in a low-salt buffer (Holbrook & Leicht,
Background
This experiment pertained to identifying an unknown bacterium. This bacterium was retrieved from a member of our group’s hand. Bacteria are prokaryotic, but like all other living organism there are many genus and species. Through experimentation these unknowns can be found can be brought to light. Bacteria can be classified either gram-positive or gram-negative. These groups differ by the composition of their cell walls. In particular, the difference in the composition pertains to amount of peptidoglycan, a polymer of amino acids and sugars, present. A Gram test is most commonly used for this classification, and pertains to using a dye to determine the classification of the bacteria. The peptidoglycan that is found …show more content…
A KOH string test is one of these methods. In this method, the bacteria is mixed with KOH, and while mixing the sample will either become viscous somewhat quickly (Positive String) or it will not (Negative String). This reaction correlates with gram-positive or gram-negative classification in a somewhat opposite manner. A positive string test indicates a gram-negative bacteria and a negative string test indicates a gram-positive bacteria. Another method that can classify gram-positive or gram-negative is the presence of bacterial growth on different media. Eosin Methlyene Blue-lactose (EMB-lactose) is an example of a media that allows gram-negative bacteria to grow and …show more content…
PCR is a common practice used to amplify a certain region of DNA. In this experiment, PCR was used to amplify the 16S ribosomal RNA gene of the bacteria. This locus was used because it is one that evolves slowly in bacteria. A Taq polymerase is used for this PCR because of the temperatures the reaction goes through. The16S rRNA primers used in this PCR, are complimentary to sequences conserved in all bacteria and will target sites of variation. These sites of variation were the regions of the DNA amplified by PCR. The products of PCR cannot be sequenced correctly without purification. Purification removes left over reactants and enzymes. A spin column-collection tube complex is used to collect the DNA during a series of centrifugation techniques with different buffers. The silica membrane holds DNA in a high-salt buffer and will release it in a low-salt buffer (Holbrook & Leicht,