Bacterial ID Lab, Lab Report Example

The virtual bacterial ID lab at http://www.hhmi.org/biointeractive/vlabs/bacterial_id took me through the steps involved in identifying a sequence of DNA. The six steps involved are sample preparation, PCR amplification, PCR purification, sequencing preparation, DNA sequencing, and sequencing analysis. Each of these steps follow exact routine procedures to ensure correct identification.

To prepare the sample, a sample must first be transferred from a culture dish into a microcentrifuge tube. Digestive enzymes are added to the sample and it is left to sit for several hours. After this, the enzymes are deactivated and the centrifuge machine is used to spin out unwanted debris. For PCR amplification, a PCR master mix solution is added to all tubes and both a positive and negative control reaction are set up. Each tube is loaded into a PCR machine that cycles through a sequence of five steps. This process allows many DNA copies to be made. In PCR purification, a microconcentrator column containing a buffer solution is put through the centrifuge machine in order to trap DNA inside the column. The column is then transferred to a second tube and put through the machine again to transfer DNA into the new tube. This separates all unwanted debris from the DNA. DNA is prepped for sequencing by adding diluted PCR product and DNA sample to many strip tubes and put through the PCR machine, this time to produce many copies of variable DNA lengths. For sequencing, the tubes are loaded into an automatic sequencer that separates the molecules based on size differences. Each nucleotide is identified.

The GenBank public database was used to make the sequencing analysis, using a matching algorithm called BLAST. After entering the sequencing code generated by the automatic sequencer, the code was automatically compared to the database. Based on the information returned from the database, I correctly identified the bacteria as Bartonella henselae.