Topic 1: The Genome and the Cell Cycle Outline PDF

Topic 1: The Genome and the Cell Cycle Lecture Objectives: 1. Describe the characteristic structures of the genome and chromosomes. 2. Describe the process of mitosis and the cell cycle, including all stages and the outcome. Focus on how the genome is organized so that each daughter cel...

Topic 1: The Genome and the Cell Cycle Lecture Objectives: 1. Describe the characteristic structures of the genome and chromosomes. 2. Describe the process of mitosis and the cell cycle, including all stages and the outcome. Focus on how the genome is organized so that each daughter cell contains a perfect copy. 3. Diagram the chemical and physical structure of DNA and understand how these structures may vary. Big Question: How do daughter cells inherit a complete copy of the genome during cell division? Topic Outline 1) The genome contains the complete set of genes typical for a species, and most cells contain a complete copy of the genome. a) Some characteristics are shared amongst prokaryotes and eukaryotes. i) Made of double-stranded DNA (dsDNA). (1) Virus genomes may be made of dsDNA, ssDNA (single stranded), dsRNA, or ssRNA, depending on the species. ii) Made of chromosomes, each of which is usually one dsDNA molecule (although this changes over the stages of cell division). Chromosomes contain: (1) Genes, which we can assume that each gene produces a product used by the cell that is necessary for cellular function. (a) Each gene is found in a particular position on a particular chromosome, thus, the complete genome will include all of these necessary genes. (2) Noncoding DNA - other (non-gene) sequences, some of which have important functions and are common to all chromosomes, such as the origin of replication sequence, which is where DNA replication begins, but others that have no known function. b) Some characteristics differ between a typical prokaryotic and eukaryotic genome: i) Prokaryotic genomes are typically: (1) Not confined within a nucleus. (2) Composed of one circular chromosome that contains one origin of replication, (3) Size ranging from 0.6 to 10 million base pairs (Mbp) (a) Size of genome is linearly related to number of genes, very little non-gene sequence. ii) Eukaryotic genomes are typically: (1) Confined within a nucleus, (2) Generally much larger than prokaryotic genomes (ie, 2.9-4,000+ Mbp). (a) Genome size does not co-vary with gene number (b) Much more non-gene sequence, which may have a function or not. (3) More likely to have repetitive sequences, meaning they are found more than once in the genome, and can be found between or even within genes. (a) Repeats are considered tandem when copies are found directly next to each other, and dispersed when they are not. (4) Made of many, linear chromosomes, each of which has at least one origin of replication. We can identify individual chromosomes in a eukaryotic genome using characteristics that are visible in karyotypes, which are images of stained chromosomes. © K. Paczolt  (a) All chromosomes have a centromere (which is an area where the chromosome shape is constricted), telomeres (ends), and chromosome arms (“p” for the short arm and “q” for the long arm). (b) Chromosomes can be identified (and named) by characteristics visible in the karyotype. These characteristics are consistent across all individuals within a species. (i) Shape, as characterized by the placement of the centromere relative to the telomeres. 1. The centromere may be at the telomere (telocentric) or progressively closer to the midpoint between telomeres. These are categorized, in order, as acrocentric, submetacentric, metacentric. (ii) Banding patterns are caused by binding of the stain (Giesma stain) to the DNA. Dark bands are areas where DNA is tightly packed, light bands are areas where DNA is loosely packed. (iii) Size. Chromosomes are named in order of descending size, so that chromosome 1 is the largest in the genome. Note: sex chromosomes (X and Y) are always placed last. (c) A haploid genome contains one copy of each chromosome, and thus one copy of each gene. (i) The haploid number (n) is the number of chromosomes found in the haploid genome. The haploid number is fixed for a species, for example, the haploid number for humans is 23. (d) Using these characteristics, we can see that most eukaryotic genomes are made up of pairs of chromosomes, called homologous chromosomes. (i) Homologous chromosomes have the same size, shape, and banding pattern, and include the same genes in the same order. 1. Within this pair, one was inherited from each parent. 2. The homologous chromosomes are not identical at the sequence level, they have small differences in sequence (SNPs). SNPs within genes may result in alleles, which are versions of a gene that affect the gene product’s function. (ii) Genomes made up of homologous pairs are called diploid genomes, meaning the genome is made up of two haploid sets of chromosomes. The diploid genome contains 2 copies of each gene. 1. A haploid set is made up of one chromosome from each pair, but, in a diploid genome, one haploid set is inherited from each parent (connect later to meiosis). 2. Each species has a typical haploid number, n, and thus a typical diploid number, 2n, which can be calculated by multiplying n by 2. 3. Haploid number and chromosomal characteristics vary between species (this will come up again in a later topic). (iii) Some variation in the chromosomes of the genome occurs within species, ie by chromosomal sex or by chromosomal disorders such as trisomy. 2) We will now examine the molecular properties of the genome. a) Genetic information is encoded in nucleic acids, which are polymers composed of nucleotide units. Characteristics include: i) The chemical composition (as discovered by Friedrich Miescher): (1) Nucleotides include three parts: (a) A pentose sugar. The carbons of the sugar are numbered, carbons 1’, 2’, 3’, and 4’ are part of the ring and the 5’ carbon is not. (i) The form of the sugar determines whether the nucleic acids is DNA or RNA. DNA uses deoxyribose sugars while RNA uses ribose sugars. © K. Paczolt  (b) The nitrogenous base, which is attached to the 1’ carbon of the sugar, and has a variable structure, which is the source of the “sequence” we think of in DNA. (i) Purines are bases with a double ring structure and include adenine and guanine. (ii) Pyrimidines are bases with a single ring structure and include cytosine, thymine, and uracil. DNA uses the thymine base while RNA uses the uracil base. (c) The phosphate group attaches to the 5’ carbon of the sugar, and has the same structure for both DNA and RNA. This group gives the nucleic acid a negative charge. (2) Chargaff discovered that in DNA the amount of guanine is always equal to the amount of cytosine and the amount of adenine is always equal to the amount of thymine. ii) The physical structure: Franklin used X-ray chromatography to find that DNA has consistent physical dimensions. iii) Watson and Crick used the chemical and physical findings together to make the double helix model of DNA, which states: (1) DNA is composed of two antiparallel strands. (a) Strands have directionality based on the carbons of the sugar. The 5’ end refers to the end of the strand with a free phosphate group and the 3’ end refers to the end with the free hydroxyl group. (2) DNA’s structure is formed so that the sugar and phosphate groups form a backbone along the outside (with nucleotides joined along a backbone by phosphodiester bonds) and the nitrogenous bases pair specifically on the inside (by hydrogen bonds). (a) In DNA, nucleotides pair based on Chargraff’s ratios, A-T and G-C, meaning that a purine always binds to a pyrimidine. As a result, DNA has a consistent width. (i) Adenine and thymine make 2 H bonds, while guanine and cytosine make 3. (ii) Areas enriched for guanine and cytosine will require more energy to separate the two strands because they have more hydrogen bonds. (iii) Strands are thus complementary – one strand can be used to predict the other – rather than identical. This will be important for DNA replication. (b) DNA winds to form a double helix. (i) Notice that the helix forms major and minor grooves. Franklin’s measurements fit the expected dimensions of the double helix as well as the width between strands. (ii) This structure is very stable, nucleotides stack on top of one another within the helix. b) DNA structure can be characterized at three levels: i) Primary structure, which is the two-dimensional chemical structure of the DNA. (1) Variation in the primary sequence of DNA includes variants in the nucleotide sequence, like SNPs (single nucleotide polymorphisms) and INDELs (insertions and deletions) as well as larger scale mutations that we will cover later. (2) The primary structure may have chemical modifications. This will be covered more later. ii) Secondary structure (1) Helices: (a) B form (or B-DNA), the double helix modeled by Watson and Crick is the most commonly found secondary structure in DNA. (b) A form (or A-DNA), occurs when dsDNA is dehydrated or in DNA-RNA hybrids. The A-DNA is a right-handed helix with 10.9 bp per turn. (c) Z form (or Z-DNA), which may form in areas with active transcription and shows a left- handed helix with 12 bp per turn. © K. Paczolt  (2) Non-helices structures may occur if a single DNA or RNA strand encodes complementary sequences. These include hairpins, stem-loops, or complex combinations and may have important functions (ie, tRNA), or not. iii) Tertiary structure, which is the folding of the double helix to further compact DNA. (1) Supercoiling occurs in both prokaryotes and eukaryotes when additional force is applied by topoisomerase enzyme to add turns to the nucleic acid, causing loops to form. (2) In eukaryotes, before supercoiling, DNA is first organized into nucleosomes. (a) Regularly spaced nucleosomes are formed by wrapping negatively-charged DNA around 8 positively-charged histones and then anchoring this shape with histone H1. (i) The cell needs many histone genes to make enough histone proteins. These are part of a class of repetitive DNA called functional repeats. 1. Usually short genes (150-300 bp length) 2. Many copies (thousands) which enables high expression of product that the cell needs many copies of 3. Often found in tandem and clustered in a few locations. 4. Other examples include rRNA and tRNA, which will be covered later. (b) Chromatin refers to the combination of DNA and bound proteins. When nucleosomes are packed close together, it forms heterochromatin, while nucleosomes not packed so tightly form euchromatin. (i) The packing of nucleosomes is mediated by the chemical structure of tails on the histones. 1. The positively charged tails usually cause chromatin to pack tightly. 2. If acetyl groups are added to the tails, they become neutral and loosely packed. (ii) It is further compacted by supercoiling (see above) which is anchored to the scaffold proteins and then then further coiled. c) Karyotype features can be recognized based on primary or tertiary structure. i) Banding Pattern: (1) Heterochromatin is the dark regions of stained DNA, and often has few genes or genes that are not highly expressed. Also, commonly found near centromeres and telomeres. (2) Euchromatin is the light regions of stained DNA, and often has many genes and/or genes that are highly expressed. ii) Telomeres have characteristic primary and secondary structures. (1) Telomeres are also made up of special tandem repetitive DNA. (a) An example of an STR, or short tandem repeat. These have short (