Chapter 10 : Gene isolation AND manipulation

A special type of nucleotide that lacks a 3' hydroxyl group, preventing further elongation of the DNA strand during replication.

A method used to determine the sequence of a DNA fragment, relying on the random incorporation of dideoxynucleotides to terminate DNA synthesis.

A continuous sequence of DNA fragments assembled from overlapping reads during genome sequencing.

A sequencing strategy that involves sequencing the entire genome in one go, rather than focusing on specific regions.

A modern sequencing technology that allows for high-throughput, parallel sequencing of millions of DNA fragments, often producing short reads.

Regions of DNA that indicate protein-binding sites, such as enzyme catalysis sites.

A gene in one organism that shares a common ancestor with a gene in another organism.

Genes arising from duplication events within the same organism.

Genes that share a common ancestor, including orthologs and paralogs.

Non-functional genes that have lost their original function due to mutations.

The process of copying and pasting genetic material within a genome, leading to the creation of new genes.

A method used to identify DNA regions bound by specific proteins by cross-linking, fragmenting, and isolating the protein-DNA complexes.

Alternative forms of mRNA produced from the same gene by splicing different exons together.

A short sequence of six histidine amino acids added to a protein, enabling purification using Ni2+-coated beads.

Circular pieces of DNA that replicate independently in bacteria, often used as vectors to carry foreign DNA.

A method for cloning larger DNA fragments, using bacteria to carry and replicate the cloned DNA.

A method for cloning large DNA fragments using phage (viruses that infect bacteria) to deliver the DNA into bacterial cells.

A method for detecting specific DNA sequences in a sample using a labeled probe that binds to the target sequence.

A technique for assembling multiple DNA fragments together, without using traditional restriction enzymes.

A method for joining DNA fragments in a specific order without leaving any unwanted sequences, useful for creating complex constructs.

The process of extracting a specific gene from an organism's DNA and creating multiple copies of it.

A specialized enzyme that cuts DNA at specific sequences, acting like molecular scissors.

A technique that amplifies specific DNA sequences in vitro using primers and a heat-stable DNA polymerase. It is used to make multiple copies of a specific gene.

Enzyme that synthesizes new DNA strands, essential for DNA replication.

A sequence of DNA recognized by a restriction enzyme, often with a symmetrical structure where the two strands read the same backward and forward.

A type of restriction enzyme cut that leaves single-stranded overhangs on the DNA fragments, allowing them to easily bind to complementary sequences.

The process of introducing foreign DNA into bacteria cells, often using plasmids as vectors, to allow the bacteria to produce the desired protein encoded by the foreign gene.

A technique used to detect protein-protein interactions. It involves fusing a bait protein with a DNA-binding domain and a target protein with an activation domain. If the proteins interact, they bring the domains together, activating gene expression.

The set of genes unique to a pathogenic strain of Escherichia coli (O157:H7) compared to a non-pathogenic strain (K12). These genes often encode virulence factors.

Proteins produced by pathogenic bacteria that contribute to their ability to cause disease. Examples include toxins, cell invasion proteins, and adherence proteins.

A region of DNA that regulates gene expression. It can be located upstream or downstream of the gene it controls, and it interacts with specific proteins to either enhance or repress transcription.

The conservation of gene expression patterns between different species. This suggests that the function of the gene is important and has been preserved through evolution.

A technique used to create cells with a specific gene knocked out. It involves using neomycin analog to kill neomycin-sensitive cells and ganciclovir to kill cells with the tk+ gene. Only cells with the targeted mutations survive.

A process where selected cells containing the targeted gene knockout are injected into blastocyst stage embryos. These embryos then develop into chimeric mice with both normal and modified cells.

Offspring of a chimeric mouse and a normal mouse. They carry a mix of genotypes and can be bred to produce homozygous strains.

The process of identifying exons and gene ends using short sequences from the 5' and 3' ends of cDNAs. It's useful for analyzing gene structure but has limitations like missing full-length sequences and multiple gene ends.

A signature sequence indicating the location of coding regions within a gene. It can be identified using cDNA sequences, RNAseq, and homology comparisons.

A technique that measures gene expression levels by converting RNA into cDNA and labeling it with primers. It detects mRNA levels, allowing for the study of gene activity.

A tool used to compare sequence similarity between genes and proteins. It identifies evolutionary relationships based on sequence alignments.

Variations in the frequency of codon usage between different species. For example, UGC and UGU both code for cysteine, but their occurrence varies between organisms.

A method of introducing genetic material into cells using viruses that have been modified to be harmless.

Directly injecting DNA into cells, often used in research with organisms like worms and mice.

A method of correcting faulty genes by using special plasmids to replace them with functional copies.

A bacterial species used to insert genes into plant cells by utilizing its Ti plasmid.

The process of creating plants with altered genetic material, typically by introducing new genes.

A method that uses homologous recombination to replace faulty genes with functional ones, resulting in a transgenic plant.

The pattern of inheritance of genes inserted using Agrobacterium tumefaciens, resulting in a 1:2:1 ratio in offspring.

The ideal outcome of homologous recombination where the targeted gene is replaced with the desired gene, while the marker gene is removed.

A possible outcome of homologous recombination where the vector DNA inserts randomly into the genome, carrying both the marker gene and the desired gene.

An outcome of homologous recombination where the vector DNA fails to insert into the genome, leaving the original genetic makeup unchanged.

A technique used to amplify specific DNA sequences by repeatedly denaturing, annealing, and extending DNA strands.

The first step in PCR, where DNA strands are separated by heating to 95°C.

The second step in PCR, where primers bind to complementary sequences on single-stranded DNA at 60°C.

The third step in PCR, where Taq polymerase adds nucleotides to the primers, extending the new DNA strand at 72°C.

A region of DNA that is recognized by a restriction enzyme, creating sticky ends.

A technique to produce RNA from a DNA template using T7 polymerase.