The DNA for a given gene in eukaryotes is organized into exons and introns. The introns do not carry any genetic information. The process of RNA splicing is responsible for removing introns from precursor RNAs to produce the final RNA product. In pre-mRNA to mRNA splicing it is critical to make sure that splicing is extremely accurate. If splicing is off by one nucleotide the entire coding will be messed up because all of the codons downstream of the mistake will be out of the correct reading frame (they will be out of phase).
RNA splicing is carried out by snRNPs which stands for small nuclear RNA containing ribonucleoprotein particles. The snRNPs contain both RNA (each snRNP contains a molecule of snRNA) and proteins. In this respect they are very similar to ribosomes, another RNP particle in the cell. In snRNPs, the RNA carries out enzymatic duties, and the proteins hold the snRNPs in the correct configuration to stabilize them.
The snRNAs in the snRNPs base pair with the pre-mRNA at splice junctions (and some other sites too). The snRNPs base paired at different splice junctions interact with each other to facilitate the removal of the intron between the snRNPs and to join the adjacent exons.
There is an evolutionary benefit to having introns; otherwise, the energy cost to splice would not be compensated.
Sometimes splicing skips over a exon. For example say the pre-mRNA
contains A-B-C-D exons. Splicing in some tissues might lead to an A-B-D mRNA (exon Cis skipped). Or the splicing could produce an A-C-D mRNA (exon B is skipped). These mRNAs would have the same end
exons but different middles. They will code for different proteins.
This alternative splicing can aid in gene regulation.
Eukaryotes differ from prokaryotes in that they:
There is great divergence of sequence between a given intron in different eukaryotic organisms. The exon sequences are much more conserved. This suggests that the actual sequence of the intron is not very important. If it were important then any changes that occurred during evolution would be damaging and the organisms with the changes would not be likely to survive.
The generally accepted solution says that the primitive genome contained no introns but consisted of short minigenes that each had very well defined functions or structures. These primitive cells also had the ability to splice out intervening sequences in the pre-mRNA. Something happened that duplicated one of the minigenes and plunked the new copy of the minigene in between two of the other minigenes. For example, if the duplicated minigene had the function of being able to bind to RNA polymerase, and it was placed next to a minigene that was capable of recognizing a specific DNA site, the new mRNA precursor would make a protein that recognizes RNA polymerase and DNA - perhaps this could be a primitive transcription factor. By plunking down this new minigene, the cell is allowed to experiment with new gene combinations and splicing without permanently altering its genome.
Processes like these might lead to evolution where minigenes were combined to form the genes as we recognize them today. Introns were the spaces between the minigenes that are now working together as a gene.
If splicing did not evolve, the duplicated minigene would have to be placed exactly touching a minigene to be able to combine functions and act as a unified gene without messing up the reading frames. The exact placement would remove any promoter upstream of the second minigene - so if the recombination "experiment" did not work the second gene becomes non-functional in the cell.
Globins (combined with heme to) bind oxygen. All globin genes have three exons and two introns. The functional protein, called hemoglobin, consists of 4 molecules of globin protein and a single molecule of heme. Human adults have two alpha- globins and two beta-globins in our hemoglobin.
Myoglobin consists of a single globin subunit plus heme and carries oxygen within muscles. Because of their similar sequence and gene organization (both have three exons in exactly the same location along the gene) it is believed that both the globin and myoglobin are derived from a common ancestor gene.
Plants called legumes have the ability to use certain kinds of bacteria as a means of getting their needed nitrogen through a process of nitrogen fixation. An example is soybeans. The roots develop a sac where bacteria can fix nitrogen. The bacteria and the plant have a symbiotic relationship; the plant provides the bacteria with food, and the bacteria fixes nitrogen for the plant. Leghemoglobin is crucial in this process because it binds oxygen within the sac which allows the bacteria to fix nitrogen. The bacteria cannot function in the presence of oxygen. The sequence of leghemoglobin is related to the sequence of the other globins, but interestingly, the middle exon is split in leghemoglobin giving this particular globin gene 4 exons. Since the gene organization is close to that of the rest of the globin family and the protein sequence of leghemoglobin and globin are related, it is clear that these genes all share a common ancestor. It is not known if the ancestor had three or four exons.
Pseudogenes are genes that do not produce a product but we believe they are genes because they have gene like qualities. They might have promoters that don't work or they may have a mutation in the coding sequence that prevents a functional protein from being produced. Pseudogenes usually arise by gene duplication of a functional gene followed by errors collecting in one of the copies.
During transformation a bacterium picks up foreign DNA which transforms the bacterium so that it now has new capabilities provided by the "foreign" gene. When the transformed bacterium replicates, the progeny will also have the new DNA. The new characteristics are heritable and stable and behave as any other part of the bacterium's DNA.
Conjugation is a process that is similar to sexual reproduction. Two bacteria cells form a conjugation bridge. The DNA from one of the bacteria moves across the bridge as it is replicated. The bridge is very fragile so many times the bridge breaks when only some of the DNA has been transferred. The transferred DNA can recombine with the original DNA (the cell with the transferred DNA is actually diploid for those genes that were transferred by conjugation). This recombination is carried out by enzymes and is similar to crossing-over in meiosis. See figure 4.2 in text.
Another way to transfer information is through phage infection. The transfer of genetic information by this means is called transduction. When the phage attacks the cell and replicates it sometimes picks up small fragments of the cell DNA and integrates the fragment into the phage head. The progeny phage will then inject this cell DNA into another cell. These recipient cells would then have a diploid situation for this DNA segment and recombination can occur.
Another mechanism by which phage can transduce DNA is to start by integrating into the host DNA through cross-over to become a prophage. Sometimes when the prophage DNA excises itself it picks up part of the host DNA with it. This extra host DNA can be delivered into the next cell the progeny phage infects resulting, again, in a partial diploid situation and produce recombination.
When behaving in this way, the phage DNA is acting as a vector. A vector DNA is any DNA that is capable of carrying a foreign piece of DNA to a different location than its source location.
Specific DNA pieces can be isolated through molecular cloning. A petri dish is covered with bacteria. Two phages are added, each of which will infect a cell. The infected cells will lyse, releasing progeny phage that will infect any nearby bacteria. The process is repeated and eventually the lysed cells will appear as a clear circle on the dish. This called a plaque which is a clear area on a bacterial lawn that contains phage. Every single progeny phage in the plaque will be identical because they all were derived from the same parent phage - all the progeny phage are clones since they contain identical genomes.
Another kind of cloning is performed with bacterial cells. One cell is placed in a petri dish. This cell will eventually form a colony of identical cells through repeated rounds of division. All of the cells will have identical information as the parent cell - these cells are all clones. See figure 4.8 in text.