BIO111 - Molecular genetics - Keywords

A repressible operon is normally expressed. Expression can be blocked in the presence of a corepressor. This corepressor is usually the product of the biochemical pathway encoded by the operon. 

Operons can be positively or negatively repressed. 

In bacteria, the DNA region to which activators and repressors bind is known as an operator. Operators usually overlap with the 3' end of the promoter and can overlap with the 5' end of the first gene in the operon. 

In gene regulation, cis and trans refer to wether mutations or gene products can affect the expression of genes only on the same DNA molecule (acting in cis) or wether they can also affect genes located on other chromosomes  (acting in trans). 

Mutations that can act in trans usually affect regulatory proteins. 

Mutations that can act in cis usually affect regulatory elements (DNA sequences) that bind regulatory proteins. 

Enhancers and silencers are regulatory elements that can control the activity of a promoter even at a great distance. They can be located either up- or downstream of the promoter. 

Enhancers increase transcription rates. 

Silencers reduce transcription rates. 

Promoter-proximal elements are elements which can be found in close proximity of the promoter. The most common of these is the TATA box, which binds TBP. Others include the CAAT and the GC box. 

Promoter-proximal elements rarely determine where a gene is expressed, but rather how strongly it is expressed. 

HAT (histone acetyl-transferase) are enzymes that acetylate conserved lysine amino acids on histone proteins. This leads to acetylated histones which results in the opening of nucleosomal structure and active chromatin. 

It is the antagonist of the histone deacetlyase (HDAc)

HDAc (histone deacetylase) is the antagonist of histone acetyl-transferase (HAT). It deacetylates the histones. This inactivates the chromatin. 

DNA methylation is a process by which methyl groups are added to DNA segments. This changes the activity of a DNA segment without changing its sequence and is known as epigenetic modification. When located in a gene promoter, DNA methylation typically acts to repress gene transcription. 

Only cytosine and adenine can be methylated. 

Epigenetics studies genetic effects not entirely encoded in the DNA sequence of an organism. 

Epigenetic inheritance describes a heritable modification in gene function / expression that is not due to changes in the base sequence in the DNA. Epigenetic states can be inherited through mitosis (and in some cases even through meiosis). 

Recombinant DNA molecules are DNA molecules formed by laboratory methods of genetic recombination. to bring together genetic material from multiple sources. This creates sequences that would otherwise not be found in the genome and is possible due to the fact that DNA molecules from all organisms share the same chemical structure. 

Application of recombinant DNA technology to solve practical problems in biology, medicine, agriculture and other areas. 

Restriction enzymes (restriction endonucleases) are DNAses expressed by bacteria to protect themselves from viruses and other foreign DNA. They recognize specific DNA sequences in viral genomes and then cut them up into pieces. Bacteria protect their own genome trough methylation of a base (often A) within the recognition sequence. Only unmethylated DNA is cleaved. 

There are 3 general types of restriction endonucleases. All of the recombinant DNA work is done with the type II enzymes. These restriction enzymes recognize short (4-8 bp), usually palindromic sequences, and cut both strands within or very close to that recognition site, leaving a 5' phosphate and a 3' OH. 

Restriction enzymes that recognize the same sequence and cut at the same position are known as isoschizomers. 

Restriction enzymes that recognize the same sequence but cut at different positions. 

A palindromic sequence is a sequence of nucleotides that, if read from 5' to 3' on one strand, matches the sequence reading from 5' to 3' on the complementary strand. 

In a blunt ended molecule, both strands terminate in a base pair. Blunt ends are not always desired in biotechnology, since when using a ligase to joint two molecules into one, the yield is significantly lower with blunt ends. 

5' and 3' overhangs are known as sticky ends, because they can base pair with complementary sequences. Sticky ends that can base pair with each other are said to have compatible ends. 

Once the base pairs have paired with each other, DNA ligase can seal the nick in the sugar-phosphate backbone, creating a covalent bond between the two. 

A clone of a gene is made by isolating and creating exact copies of just one gene of an organism. This involves copying the DNA sequence of that gene into a smaller, more easily manipulated piece of DNA, such as a plasmid. 

Cloning is the process of creating an geneticly identical replica of a donor organism or cell. 

A cloning vector is a stable, self-replicating DNA molecule to which a foreign DNA fragment can be attached for introduction into and replication in a host cell. 

An effective vector has three characteristics: 

1. An origin of replication, which allows replication in the host cell

2. One or more selectable markers, which allow the selective growth or identification of cells that have taken up the vector

3. One ore more unique sites, into which the foreign DNA can be inserted

The most common bacterial cloning vectors are plasmids. 

Plasmids are small, circular extrachromosomal DNA molecules present in many bacterial species. In the wild, plasmids often carry genes that are not essential for bacterial function, but that can play important roles in the life cycle or growth of bacteria. Their sizes range from a few kb to a few hundred kb, and they have their own origin of replication (ORI), replicating independently of the bacterial chromosome. 

They are further differentiated by the number of their copies present in the cell. Stringent plasmids are present only in one or two copies while relaxed plasmids can be present up to many dozen times. 

A cosmid is a type of hybrid plasmid. It contains a Lambda phage cos sequence and is often used as a cloning vecor in genetic engineering. 

Reverse transcriptase is an enzyme that basically does the opposite of transcription, by making a DNA copy from a RNA template. This is done because a bacteria can't process introns. In order to remove the introns, mRNA (where the introns have already been spliced out) is copied by reverse transcriptase. A cDNA (copyDNA) is created, which now consists only of exons, and which can be incorporated into a vector. 

cDNA or copy DNA is a DNA copy of an mRNA template. 

PCR is an in vitro amplication of DNA fragments.

It requires a ssDNA template, a primer, a DNA polymerase and dNTPs.

It consists of many cycles of DNA replication. In each cycle, only the region of interest is replicated, as only the oligonucleotides added by the biologist can be used as primers by the DNA polymerase. 

ddNTPs are nucleotides used for sequencing. The trick is, that they lack a -OH group on both 2' and 3' carbon atoms. This means that they can be added to a growing nucleotidechain by DNA polymerase, but they can not form a bond with the next dNTP. DNA polymerase thus stops DNA polymerization as sson as a ddNTP is added to a growing chain. ddNTPs are therefore also known as chain terminators.