Genes, Genomes, and Chromosomes: Structure, Organization, and Analysis

Genes, Genomes, and Chromosomes

Introduction

This chapter explores the structure, organization, and analysis of genetic material in prokaryotes, eukaryotes, and viruses. It covers the physical packaging of DNA, the differences between chromatin and chromosomes, and modern biochemical tools for genome analysis.

21.1 Bacterial and Viral Genomes

Fundamental Terms of Genetic Information

• Gene: A chromosomal segment that encodes a single polypeptide chain, an RNA molecule, or has a regulatory function.

• Genome: The total genetic information contained in a cell, organism, or virus.

• Chromatin: DNA complexed with histones and other proteins, typically dispersed throughout the nucleus during interphase.

• Chromosome: A compact strand of DNA encoded with genes, composed of chromatin, and formed during nuclear division.

Properties of Bacterial and Viral Genomes

Bacterial and viral genomes vary in size, gene number, and physical structure. The following table summarizes key properties:

Organism or Virus	Genome Size (bp or bases)	Number of Genes	Physical Nature of Genome
Escherichia coli	4,639,221	~4400	Circular Duplex
Bacteriophage T4	168,889	~175	Linear Duplex, Circularly Permuted
Bacteriophage T7	39,936	~35	Linear Duplex, small repeat at each end
Bacteriophage λ	48,502	~50	Linear Duplex, single stranded ends
Mimivirus	1,181,404	979	Linear Duplex
Influenza Virus	~13,500	12	Single-stranded RNA

Additional info: Circular permutation means all genomes have the same linear sequence of genes, but the beginning and ending points vary among different genomes.

Packaging of Bacterial DNA

• Bacterial DNA is compacted by negative supercoiling.

• This compact structure is called a nucleoid, which exists in the cytosol with some membrane attachments.

21.2 Eukaryotic Genomes

Comparison of Prokaryotic and Eukaryotic Genomes

• Eukaryotic genomes are generally much larger than prokaryotic genomes.

• Bacteria typically have a single chromosome per cell, while eukaryotes usually have two copies of each chromosome (diploid), except for sex chromosomes (e.g., X and Y in males).

21.3 Physical Organization of Eukaryotic Genes: Chromosomes and Chromatin

The Nucleus

• The nuclear envelope contains pores (~9 μm diameter) for diffusion of small molecules between nucleus and cytoplasm.

• Exportins and importins mediate selective transport of RNA and proteins.

• The nuclear pore complex is a large assembly (500–1000 proteins, called nucleoporins).

• Transcription (in nucleus) and translation (in cytoplasm) are separated in eukaryotes.

Chromosomes

• Chromosomes are visible during metaphase as replicated structures with two chromatids joined at the centromere.

• Telomeres are DNA repeat sequences at chromosome ends, protecting DNA from degradation and ensuring complete replication.

Chromatin Structure

• The total length of DNA in a human cell is about 2 meters, but it fits in a nucleus ~10 μm in diameter.

• DNA is wound around histone and nonhistone proteins to form chromatin.

• Nucleosomes are the fundamental units of chromatin, containing 146 bp of DNA wrapped around a histone octamer (2 each of H2A, H2B, H3, H4).

Structure and Properties of the Nucleosome

• DNA is complexed with histones: H2A (yellow), H2B (red), H3 (blue), H4 (green).

• Nucleosomes are revealed by X-ray diffraction as core particles.

Higher Order Chromatin Structure

• Chromatin during interphase exists as:

  • Euchromatin: Transcriptionally active, less condensed.

  • Heterochromatin: Transcriptionally inactive, more condensed.

• Chromosome conformation capture experiments show that chromatin, though appearing disorganized, has a higher degree of order.

• Chromosome neighborhoods are regions of DNA that are far apart linearly but close together in 3D space.

21.4 Nucleotide Sequence Analysis of Genomes

Sequencing a Genome

• Shotgun sequencing generates random DNA fragments; overlapping regions are used to assemble the genome sequence.

• The Human Genome Project (1990–2003) sequenced the 3 billion base pairs of the human genome using advanced techniques.

Mapping Large Genomes with Fluorescent In Situ Hybridization (FISH)

• FISH identifies the location of a gene on a chromosome by tagging DNA with a fluorescent dye.

• The DNA probe is denatured and annealed to chromosomes; the dye's location is visualized by microscopy.

DNA Fingerprinting

• Uses restriction fragment length polymorphisms (RFLPs) to distinguish individuals based on DNA patterns.

• PCR can amplify small amounts of DNA for analysis.

• Most RFLPs are due to changes in short tandem repeats, not single base changes.

Size of the Human Genome

• The human genome contains about 3 billion base pairs and 20,000–25,000 genes.

• ENCODE project results suggest noncoding DNA has important functions.

• Any two humans differ at about 1 base per 1,000 base pairs.

21.5 Tools of Biochemistry

Polymerase Chain Reaction (PCR)

• Invented by Kary Mullis in 1983, PCR revolutionized molecular biology by enabling exponential amplification of DNA in vitro.

• Requirements: thermostable DNA polymerase (e.g., Taq), oligonucleotide primers, dNTPs, and a DNA template.

Applications of PCR

• Forensics: Amplification of DNA from biological samples (blood, semen, hair).

• Cloning: Amplification of specific DNA sequences for insertion into vectors.

• Quantitative PCR (qPCR): Assessing abundance of target DNA.

• Reverse transcription PCR (RT-PCR): Monitoring gene expression by analyzing mRNA levels.

Summary Table: Key Differences Between Prokaryotic and Eukaryotic Genomes

Feature	Prokaryotes	Eukaryotes
Genome Size	Small (thousands to millions of bp)	Large (millions to billions of bp)
Chromosome Number	Usually one	Multiple, usually diploid
DNA Packaging	Supercoiling, nucleoid	Chromatin, nucleosomes, higher order structures
Location	Cytoplasm	Nucleus