Readings for Monday's class

On Monday we will start with chapter 4. Read pages 195-201!

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LAB - GMO gel electrophoresis

In this lab we are simply running the gels to visualize the results of our GMO-testing PCRs. We prepared 2.5% agarose gels as opposed to the most common 1% gels, because the expected fragments are small and very close in size (CMV 35S: ~200 bp; NOS: ~220 bp). In cases like these a denser gel will increase the resolution of the gel by enhancing band separation and making sharper bands.

There were six samples in our gels

Lane 1: Certified non-GMO plant DNA - Plant primers

Lane 2: Certified non-GMO plant DNA - GMO primers

Lane 3: Corn or soy DNA - plant primers

Lane 4: Corn or soy DNA - GMO primers

Lane 5: Positive control DNA - plant primers

Lane 6: Positive control DNA - GMO primers

Lane 7: Ladder

NOTE: Do not worry about questions 2 and 3 on pages 43 and 44 of lab 6 (question 3 is not actually labeled, so skip to question 4).

Here are two of the gels showing the best results. If for some reason your gel didn't come out well, or your PCRs didn't work write that down in your results and use one of these images to compare with your results in the discussion section.

Gel in section 01

Gel in section 02

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LAB - Genetically Modified Organisms (GMOs)

Bio-Rad's GMO Investigator Kit

In this lab we are trying to find evidence of genes that have been inserted into a plant's genome. There are two ways of finding out:

1. Testing for the presence of proteins that the trans-genes produce
   
2. Amplifying a fragment of DNA that contains the trans-gene via PCR
   

Since we need a lot of fresh tissue and a kit to test for several of the proteins that the most common trans-genes code for, materials that we don't have, we are using the almighty PCR.

Most trans-genes have the 35S promoter from the cauliflower mosaic virus (CMV 35S), and the nopaline synthase (NOS) terminator from Agrobacterium tumefaciens. These sequences come from totally different organisms from that used to isolate the actual trans-gene, but they are recognized by the plant cell machinery (mainly its polymerase), so they can actually produce the product that the gene is intended for. Even if the gene is in the genome, it will NOT work if there are no promoters AND terminators that the plant can recognize. Since there is a number of genes that have been inserted in crops in the U.S., it's easier to go after the promoter and the terminator when using PCR. CMV 35S and NOS have been use in over 85% of the GMOs, so if we have primers to amplify them, we don't have to worry about which gene was inserted (as long as the question doesn't involve knowing the specific gene).

We extracted DNA from corn or soy that we presume have been genetically modified, and from a cerified non-GMO food control provided by Bio Rad, the manufacturers of the kit we use. The master mixes we used include primers for both CMV35S and NOS in the same mix, and in addition to those we are using a separate master mix with primers that will amplify a 455 bp region of the photosystem II (PSII) chloroplast gene, common to most plants. This will be a control to ensure we have successfully extracted DNA, so even non-GM plants should have the band present.

Speaking of PCR, watch the Bio Rad PCR animation NOW...!

Check the Bio-Rad web page for this lab!

The gel electrophoresis will be ran the following day.

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Genomics and the tree of life

Click on the pic for a full-size image

Today we talked about the diversity of genomes and thier role in recostructing the tree of life. The material is basically covered on pages 15-24 of Alberts' book.

Goals of the topic

1. Understand the main reasons for the success of DNA-based life
2. Get a basic understanding of how genomes evolve
3. Understand how features of genomic evolution shape the tree od life
4. Understand the impact of genomics in the field of systematics
   

We well resume class starting at "New genes are generated from preexisting ones" (p 18).


Quiz #3 - Q&As

1. What is a codon?
In a gene, a series of three nucleotides that encode information that will be ultimately translated into an amino acid in a protein

2. What is translation?
The process in which proteins are synthesized according to RNA sequences carrying over infromation from the DNA

3. What is transcription?
The process in which information found in the DNA is passed onto an RNA molecule

4. What are the three domains in the tree of life?
Archea (Archebacteria), Bacteria (Eubacteria), and Eukaryota

5. What is a gene family?
A group of genes in a genome that descend (via gene duplication) from a single gene

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