Genetics Ch 20

The 2 important genes vector carries

1. ampR 2. lacZ

Which component in the CRISPR-Cas9 system makes a double-strand break in DNA?

Cas9

Genes are cloned by splicing a fragment of _____ from an organism into a cloning vector to make a recombinant DNA molecule

DNA

A collection of recombinant vectors

DNA library

A researcher may use restriction enzymes to digest the DNA of an organism. The fragments of DNA are then ligated individually into many vectors. This collection of recombinant vector is called a ______ ______

DNA library

After ligation, there is a collection of recombinant vectors, with each vector containing a particular fragment known as a ____ ______

DNA library

DNA sequencing enables reseacrhers to determine the order of ______ _____ in a gene

DNA nucleotides

Dideoxy sequencing was formulated based on scientist knowledge of what process?

DNA replication

This technique enables researchers to determine the DNA bases in genes and other chromosomal regions

DNA sequencing

The DNase I footprinting technique is used to study _______-_____ interactions

DNA-protein

The most common plasmids are the ___ ____ plasmid cloning vectors

E. coli

Director of Human Genome Project

Francis Collins

Which scientist developed the polymerase chain reaction?

Kary Mullis

If we want only one region of genomic DNA or one mRNA then we use _______ ________ _______

Polymerase Chain Reaction

_______ _____ occurs when short oligonucleotides bind to complementary DNA flanking the gene of interest

Primer annealing

____ _____ _____ is used to quantitate the amount of a specific Target DNA

Real-Time PCR

The use of in vitro molecular techniques to isolate and manipulate fragments of DNA

Recombinant DNA technology

Cuts the phosphate-sugar backbone between the phosphate and the 3' carbon

Restriction Enzyme

Multiple cloning site = ______ ______ _____

Restriction site cluster

____-______ ________ alows a reseacher to produce a mutation at a specific sequence

Site-directed mutagenesis

In PCR, each cycle uses the products of the previous cycle as a template. What do you call this?

a chain reaction

In PCR, each cycle uses the products of the previous cycle as templates. What do you call this?

a chain reaction

Confers antibiotic resistance to the host cell and identifies cells that have taken up the vector

ampR

If the starting material was cDNA, the library is called a _____ _______

cDNA library

An actual ______ library would contain thousands of different bacterial colonies. Each one carrying a different piece of cDNA created from the starting _____ sample

cDNA; RNA

Oligonucleotides called linkers, with a restriction enzyme site, are attached to the ______ using ____ _______

cDNAs; DNA ligase

A particular gene to be cloned is often isolated from ______ _____

chromosomal DNA

To make many copies of a gene, you would _____ that gene

clone

A DNA molecule that acts as a carrier of DNA that is to be cloned

cloning vector

A molecule that carries the DNA to be cloned

cloning vector

Cells that are able to take up DNA

competent cells

In PCR, primer extension refers to the synthesis of _________ _____ starting at the primers

complementary DNA

The DNA sequencing method developed by Frederick Sanger that became a commonly used method of DNA sequencing

dideoxy sequencing

The gel retardation assay is also know as the ______ ____ _____ _____

electrophoretic mobility shift assay

What technique is useful for studying protein-DNA interactions?

electrophoretic mobility shift assay and DNase I footprinting

During the initial phase of a real-time PCR experiment, called the _________ phase, the amount of PCR products is small and reagents are not limiting, so the amount of product nearly doubles with each cycle

exponential

The amount of product nearly doubles with each cycle but may be difficult to detect as the amounts are small

exponential

In automated sequencing, each dideoxyribonucleotide is labeled with a different-colored _______ _____

fluorescent dye

The TaqMan system uses a detector oligonucleotide that has a _______ _______ _______ at one end and a ________ ______ at the other end

fluorescent reporter molecule; quencher molecule

The process that produce many copies of a DNA moledcule of interest

gene cloning

If the starting material was chromosomal DNA, the library is called a ______ _______

genomic library

An actual ______ library would contain thousands of different bacterial colonies. Each one carrying a different piece of ________ _____

genomic; chromosomal DNA

Very active Ori, generates a _____ copy number per cell

high

A cell that harbors a vector

host cell

Recombinant DNA molecule is introduced into a _____ _____ where it is replicated aka ________

host cell; transformation

Encodes β-galactosidase and provides a means by which bacteria that have picked up the cloned gene can be identified

lacZ

In the hybrid vector, the chromosomal DNA inserts into the ____ gene, thereby disrupting it

lacZ

Accumulation of the product shows a directly proportional relationship to cycle number

linear

In PCR, the temperature must be _______ from the denaturation temperature in order for primers to anneal

lowered

This process indentifies a specific RNA moledcule within a mixture of RNA molecules

northern blotting

Human chromosomal DNA generate thousands of fragments. Typically ____ fragment inserts into a vector and becomes a ________ _______

one; recombinant vector

Allows for replication of plasmid in E. coli

ori sequence

A vector requires an _____ ___ ______ so that it can be copied many times by the host cell

origin of replication

What do you call the DNA sequence in a vector that allows that replication enzymes of the cell to make lots of copies of the vector?

origin of replication

________ means the base sequence reads the same backwards and forwards.

palindromic

A small circular DNA moledcule that is often used as a vector in gene cloning

plasmid

Transformation refers to uptake of _______ vectors by a ______ cell

plasmid; bacterial

Plasmids used for cloning experiments are derived from naturally occurring ______ that have been modified to facilitate cloning

plasmids

Accumulation of the product level off as one or more reagents are used up

plateau

In 1985, Kary Mullis developed a way to copy DNA without vector or host cells. This technique is called _______ ______ _____

polymerase chain reaction

During PCR, the process of _______ ______ results when the Taq polymerase catalyzes the synthesis of complementary DNA, starting at the primers

primer extension

Short oligonucleotides that flank the region of DNA to be amplifies by PCR

primers

When using PCR to amplify DNA, short oligonucleotides called ________ flank the region of DNA to be amplified

primers

What is the technique that allows one to determine the amunt of template DNA present when the PCR cycles began?

quantitative PCR

_______-_____ PCR allows one to assess the amount of DNA produced during a PCR amplification as it is happening

real-time

In contrast to the hybrid vector, the _________ vector has a functional ____ gene

recircularized; lacZ

The production of new arrangements of DNA

recombinant DNA technology

The use of in vitro molecular techniques that combine DNA fragments to produce novel arrangements

recombinant DNA technology

A vector that contain a piece of chromosomal DNA

recombinant vector

Vector that contains a piece of chromosomal DNA

recombinant vector

ampR

resistance to ampicillin

camR

resistance to chloramphenicol

tetR

resistance to tetracyclin

Enzymes that bind to a specific DNA sequence and cut the DNA backbone

restriction endonucleases

_______ ______ are used in gene cloning to cut the DNA backbone prior to inserting the DNA to be cloned

restriction endonucleases

Site-directed mutagenesis allow a researcher to make a mutation at a ________ _____ of DNA

specific sequence

In PCR, the DNA to be amplified is called the _______ DNA

template

In PCR, the _____ DNA is the DNA to be amplified

template

Treating chromosomal DNA with restriction enzymes can generate _____ ___ _______ of fragments

tens of thousands

When cloning a gene, why must the chromosomal DNA and the plasmid DNA be cut with the same restriction enzyme?

the sticky ends of the plasmid DNA will be complementary to the sticky ends of the chromosomal DNA

Use a ______ _________ (essential equipment) to generate different temperatures for each cycle

thermal cylcer

True or False: Chromosomal DNA is a common source of cloned DNA

true

A small DNA molecule that can replicate independently within a host cell

vector

A small DNA molecule that can replicate independently within a host cell and thus make many copies of an inserted gene

vector

Transfection refers to introduction of ______ vectors into a ____ _____

viral; host cell

What technique is used to identify a particular protein in a mixture of proteins?

western blotting

You wish to determine if a protein is made at a particular stage of development. What technique would you use?

western blotting

An E. Coli Cloning Vector Must Have Three Features

1. Ori sequence 2. Selectable marker 3. restriction enzyme cleavage sites

The enzyme _______ ________ is used when PCR is emplyed to detect and quantify the amount of a specific RNA

reverse transcriptase

The enzyme that uses RNA as a template to make a complementary strand of DNA

reverse transcriptase

You have a piece of RNA, and you want to synthesize a complementary strand of DNA. Which enzyme would you use?

reverse transcriptase

_______ ______ PCR can be used to detect and quantify the amount of specific RNA

reverse transcriptase

The use of dideoxyriblonucleotides with different colored fluorescent dyes allows the detection of the _______ ___ ____

sequence of DNA

To perform PCR, a machine called a _________ automated the timing of each cycle

thermocycler