Smart Book 20

Order the following steps in cloning a gene, putting the first step at the top.

1. Chromosomal DNA is isolated and cut with a restriction enzyme; the plasmid DNA is cut with the same enzyme. 2. The digested chromosomal DNA and plasmid DNA are incubated together. 3. Ligation by DNA ligase.

What does the enzyme DNA ligase do?

It covalently links the sugar-phosphate backbone of DNA.

Why would you use a poly-dT primer when making cDNA?

It would be complementary to the poly-A tail at the 3' end of the mRNA.

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

Western blotting

Why is Taq polymerase used in PCR?

The DNA polymerase must be thermostable as PCR involves cycles of heating.

DNA sequencing enables researchers to determine the order of ______ ______ in a gene.

DNA nucleotides

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

vector

Order the steps in one cycle of a PCR reaction, putting the first step at the top.

1. Denaturation 2. Primer annealing 3. Primer extension

In dideoxyribonucleotides, ____ oxygens are removed from the sugar compared with ribose.

2

Chain termination occurs when a dideoxyribonucleotide is incorporated into a growing DNA strand because there is no ______.

3'-OH group

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

A chain reaction

In gene cloning, what is the vector?

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

What is a plasmid?

A small circular DNA molecule often used as a vector in gene cloning

In PCR, the two primers bind to specific sites in the ____ and flank the gene to be amplified.

DNA

Dideoxy sequencing was formulated based on scientists' knowledge of what process?

DNA replication

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

DNA sequencing

Which scientist developed the polymerase chain reaction?

Kary Mullis

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

Polymerase chain reaction (PCR)

How can PCR amplify one segment of DNA from a complex mixture of potential template molecules?

Primers can be designed to flank a specific segment of DNA.

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

Reverse transcriptase

What is recombinant DNA technology?

The production of new arrangements of DNA

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.

In PCR, why do the primers bind to specific sites in the DNA on either side of the gene of interest?

They are complementary to the flanking sequences.

What is the purpose of gene cloning?

To produce many copies of a DNA molecule of interest

True or false: Amplifying a gene by PCR results in many copies, just like cloning using a vector and host cell.

True

True or false: Chromosomal DNA is a common source of cloned DNA.

True

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

Western blotting

You would ____ a gene to make many copies of that gene.

amplify

A recombinant DNA molecule has covalently linked DNA fragments from ______.

at least two different sources

In dideoxy sequencing, if a dideoxyribonucleotide is incorporated into the growing strand of DNA, the strand can no longer grow as there is no 3' OH group. This is called ______.

chain termination

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

chromosomal DNA

If a gene is amplified by PCR so that there are many copies, it can be said to be ______.

cloned

Making many copies of a particular DNA segment using vectors or the polymerase chain reaction is called gene

cloning

Transformation occurs when ______.

competent cells take up DNA from the medium

Restriction endonucleases are used in gene cloning to ______.

cut the DNA backbone prior to inserting the DNA to be cloned

The DNA sequencing method developed by Frederick Sanger that became a commonly used method of DNA sequencing is called ____ sequencing.

dideoxy

If the oxygens on carbons 2 and 3 of the sugar of a nucleotide have been removed, the nucleotide is referred to as a

dideoxyribonucleotide, ddNTP

The replication of recombinant DNA molecules inside a host cell is one form of ______.

gene cloning

When cloning a gene into a vector, the sugar-phosphate backbone of each DNA molecule is covalently linked by the enzyme DNA

ligase

Select the reagents needed to make cDNA.

mRNA Poly-dT primer dNTPs Reverse transcriptase

A small circular DNA molecule that is often used as a vector in gene cloning is called a(n)

plasmids

____ DNA technology uses in vitro molecular techniques that combine DNA fragments to produce novel arrangements.

recombinant

A molecule that has covalently linked DNA fragments from at least two sources is called a ____ ____ molecule.

recombinant DNA

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

restriction enzymes

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

reverse transcriptase

In PCR, the DNA to be amplified is called the ____ DNA.

template

In PCR, the template DNA is ______.

the DNA to be amplified

Taq polymerase was first isolated from a bacterium called

thermus aquaticus

The process by which competent cells take up DNA from the extracellular medium is called

transformation

True or false: PCR can amplify one segment of DNA from a mixture. True false question.

True