Topic 8 Producing DNA Fragements

Question 1

What is Recombinant DNA Technology?

Why is it so important?

Answer 1

Recombinant DNA Technology is combining of different organisms DNA.

It is important as it enables scientists to manipulate and alter genes to improve industrial processes and medical treatment.

Question 2

What is the first step in Recombinant DNA Technology?

Answer 2

First step is to produce or isolate the fragments of DNA

To be recombined with another piece of DNA

Question 3

What are the three methods in creating DNA Fragments?

Answer 3

1. Reverse Transcription

2.Restriction Endonucleases

3.Gene Machine

Question 4

What occurs in Reverse Transcription and where does it occur?

What is the first step in in Reverse Transcription?

Answer 4

In Reverse Transcription reverse transcriptase makes DNA copies from mRNA which naturally occurs in viruses such as HIV

First step in a cell naturally produces protein of interest is selected.

Question 5

What is selected in Reverse Transcription and what must it contain?

Answer 5

A Cell naturally produces the protein of interest is selected.

One of the cell should have large amounts of mRNA for protein

Question 6

What is the function of Reverse Transcriptase Enzyme?

Answer 6

Reverse Transcriptase Enzyme joins the DNA nucleotides with complementary bases to mRNA sequence to make single stranded DNA (cDNA)

Question 7

What is cDNA?

Answer 7

Single Stranded DNA

Question 8

In Reverse Transcription what is needed to make DNA fragments double stranded?

Answer 8

Enzyme DNA Polymerase is used to make double stranded DNA fragments.

Question 9

What is the advantage of cDNA?

Answer 9

cDNA is intron free

Because based on mRNA sequence

Question 10

What occurs in Restriction Endonucleases?

Where does Restriction Endonucleases naturally occur?

Answer 10

Restriction Endonucleases are enzymes that cut up DNA

Restriction Endonucleases naturally occur in Bacteria

Question 11

What makes Restriction Endonucleases to cut DNA?

Answer 11

Restriction Enzyme that have an Active Site Complementary in shape to different DNA base sequences which is described as Recognition Sequences

So each enzyme cuts the DNA at specific location

Question 12

How is a blunt end is created in Restriction Endonucleases?

Answer 12

Restriction Enzymes cut at the same location in double strand.

Question 13

If blunt end is not created in producing DNA fragments what is made and how is it made?

Answer 13

Restriction Enzymes cut to create staggered ends and exposed DNA bases

Question 14

Described staggered ends and what is the advantage of staggered ends

Answer 14

Staggered ends are palindromic and referred to as ‘sticky ends’

‘Sticky ends’ on DNA fragments make it easier to insert into organism to make Recombinant DNA

Question 15

What is the disadvantage of Restriction Endonucleases?

Answer 15

Still contains introns

Question 16

What is the Gene Machine?

Answer 16

Gene Machine produces DNA fragments and is created in lab using computerised Machine

Question 17

What is the first two steps in Gene Machine?

Answer 17

1.Scientist first examine the protein of interest to identify the AminoAcid sequence and from that what the mRNA AND DNA sequence would be

2.(DNA sequence) entered into the computers which checks for bio safety and bio security which checks if DNA is safe and ethical to produce.

Question 18

So how does Gene Machine create a gene?

Answer 18

Computer create Small Section of overlapping single strands of nucleotide that make gene called

Oligonucleotides

Oligonucleotides can be joined to create the DNA for entire gene

Question 19

What is and how is Oligonucleotides made?

Answer 19

Small section opt overlapping Single Strands of nucleotides that make Oligonucleotides

Question 20

How is double stands produced in Gene Machine?

What is the advantage?

Answer 20

PCR can be used to Amplify the quantity and make the double stand

Advantage: very quick and accurate process and makes introns free

Question 21

What is the disadvantage and the advantage of Gene Machine?

Answer 21

Disadvantage: Need to know the sequence of AminoAcid or base

Advantage: Can design exact DNA fragments you want with sticky ends, labels and preferential codons

Question 22

What is the advantage of Reverse Transcriptase and the disadvantage?

Answer 22

Advantage: mRNA present in cells is actively transcribed genes so lots of the mRNA of interest available to make cDNA

Disadvantage: More steps so more time consuming so technically different