Genes and Chromosomes

Chromosomes are the carriers of genes.

KEY TERMS: gene, allele, dominant, recessive, genotype, phenotype, chromosome, diploid, meiosis, haploid, mutation, recombination, genetic map

Genes and Enzymes

A gene specifies the amino acid sequence of a polypeptide chain.

KEY TERMS: one gene-one enzyme hypothesis

Identification of DNA as the Genetic Material

DNA was identified as the genetic material by bacterial transformation experiments.

KEY TERMS: transformation

The Structure of DNA

DNA is a double helix in which hydrogen bonds form between purines and pyrimidines on opposite strands. Because of specific base pairing—A with T and G with C—the two strands of a DNA molecule are complementary in sequence.

Replication of DNA

DNA replicates by semiconservative replication, in which the two strands separate and each serves as a template for synthesis of a new progeny strand.

KEY TERMS: semiconservative replication, DNA polymerase

Colinearity of Genes and Proteins

The order of nucleotides in DNA specifies the order of amino acids in proteins.

The Role of Messenger RNA

Messenger RNA functions as an intermediate to convey information from DNA to the ribosomes, where it serves as a template for protein synthesis.

KEY TERMS: central dogma, transcription, translation, messenger RNA (mRNA), RNA polymerase, ribosomal RNA (rRNA), transfer RNA (tRNA)

The Genetic Code

Transfer RNAs serve as adaptors between amino acids and mRNA during translation. Each amino acid is specified by a codon consisting of three nucleotides.

KEY TERMS: genetic code, in vitro translation, codon

RNA Viruses and Reverse Transcription

DNA can be synthesized from RNA templates, as first discovered in retroviruses.

KEY TERMS: retrovirus, reverse transcription, reverse transcriptase

Restriction Endonucleases

Restriction endonucleases cleave specific DNA sequences, yielding defined fragments of DNA molecules.

KEY TERMS: restriction endonuclease, gel electrophoresis, restriction map

Generation of Recombinant DNA Molecules

Recombinant DNA molecules consist of a DNA fragment of interest ligated to a vector that is able to replicate independently in an appropriate host cell.

KEY TERMS: molecular cloning, vector, recombinant molecule, molecular clone, DNA ligase, cDNA

Vectors for Recombinant DNA

A variety of vectors are used to clone different sizes of DNA fragments.

KEY TERMS: plasmid, origin of replication, cosmid, yeast artificial chromosome (YAC)

DNA Sequencing

The nucleotide sequences of cloned DNA fragments can be readily determined.

KEY TERMS: dideoxynucleotide, autoradiography

Expression of Cloned Genes

The proteins encoded by cloned genes can be expressed at high levels in either bacteria or eukaryotic cells.

KEY TERMS: expression vector, baculovirus

Amplification of DNA by the Polymerase Chain Reaction

PCR allows the amplification and isolation of specific fragments of DNA in vitro.

KEY TERMS: polymerase chain reaction (PCR)

Nucleic Acid Hybridization

Nucleic acid hybridization allows the detection of specific DNA or RNA sequences.

KEY TERMS: nucleic acid hybridization, probe, Southern blotting, Northern blotting, in situ hybridization

Detection of Small Amounts of DNA or RNA by PCR

PCR provides a sensitive method for detecting small amounts of specific DNA or RNA molecules.

Antibodies as Probes for Proteins

Antibodies are used to detect specific proteins in cells or cell extracts.

KEY TERMS: antibody, antigen, monoclonal antibody, immunoblotting, Western blotting, immunoprecipitation, SDS-polyacrylamide gel electrophoresis (SDS-PAGE)

Probes for Screening Recombinant DNA Libraries

Specific DNA inserts can be detected in recombinant DNA libraries by the use of either nucleic acid hybridization or antibody probes.

KEY TERMS: recombinant DNA library, genomic library, cDNA library

Genetic Analysis in Yeasts

The simple genetics and rapid replication of yeasts facilitate the molecular cloning of a gene corresponding to any yeast mutation.

KEY TERMS: temperature-sensitive mutant

Gene Transfer in Plants and Animals

Cloned genes can be introduced into complex eukaryotic cells and multicellular organisms for functional analysis.

KEY TERMS: gene transfer, transfection, transient expression, liposome, electroporation, transgenic mouse, embryonal stem (ES) cell, Ti plasmid

Mutagenesis of Cloned DNAs

In vitro mutagenesis of cloned DNAs is used to study the effect of engineered mutations on gene function.

KEY TERMS: reverse genetics, in vitro mutagenesis

Introducing Mutations into Cellular Genes

Mutations can be introduced into chromosomal gene copies by homologous recombination with cloned DNA sequences. In addition, the expression or function of specific gene products can be blocked by antisense nucleic acids or dominant inhibitory mutants.Chapter 3

KEY TERMS: homologous recombination, antisense nucleic acids, dominant inhibitory mutant