recombinant DNA technology

Question 1

what does DNA recombinant technology involve?

Answer 1

the transfer of fragments of DNA from one organism or species, to another

Question 2

Give na application of DNA recombinant technology

Answer 2

allow large quantities of pure proteins to be producde

Question 3

give the 5 steps of using DNA recombinant technology

Answer 3

• isolation/ production
• insertion
• transformation
• identification
• growth/cloning

Question 4

Describe the 5 steps of using DNA recombinant technology

Answer 4

• isolation/ production of DNA fragments that contaion the desired gene
• insertion opf the DNA fragment into a host cell
• transformation ie the transfer f DNA into suitable host cells
• identification of the host cells that have taken up the gene
• growth/cloning of the population of host cells

Question 5

what makes obtaining fragments of DNA potentially difficult?

Answer 5

must find the gene then isolate it from millions of beases making up the gene

Question 6

give some othe methods of obtaining DNA fragments

Answer 6

• enzymes caklled restriction endonucleases to cut fragments
• converting mRNA to cDNA using recerse transciptase
• creating the gene in a gene machine

Question 7

define the term sticky ends

Answer 7

• single stranded sections of DNA
• they form an overhang at the end of a double stranded molecule
• these single stranded bases are exposed and can ben to a complementarty sticky end

Question 8

decrivbe how do restriction endonucleases work

Answer 8

• cuts dna at a specific sequence of bases

- by breaking the phosphodiester bonds in the sugar phosphate backbone

Question 9

what does palindromic mean?

Answer 9

the base sequenmce on one strand is the same sequence as on the other strand, but in reverse

Question 10

Suggest two advantages of using reverse transcriptase to produce a gene in comparison to removing DNA from an organism’s genome using restriction endonucleases.

Answer 10

• mRNA is at a high concentration within cells therefore it easier to isolate compared to single DNA copy of a gene
• The cDNA produced will have the non-coding introns removed. The gene will therefore be able to be expressed in prokaryotes

Question 11

why is the new molecule called cDNA when using reverse transcriptase?

Answer 11

the DNA molecule produced is complementary to the original mRNA molecule

Question 12

What is the function of the DNA polymerase?

Answer 12

• It synthesises a second DNA strand using the single stranded cDNA as a template, to form double-stranded DNA.
• It forms the bonds that create the sugar phosphate backbone, joining the new nucleotides together.

Question 13

If attempting to obtain the gene that codes for the hormone insulin, why
must pancreatic cells be used for this technique?

Answer 13

Only transcribe mRNA in these cells

Question 14

What could be an advantage of producing artificial genes in this manner ie gene machine?

Answer 14

-The genes would be free of introns and other ‘non-coding’ DNA (so they can
transcribed and translated by prokaryotic cells)
-any sequence of nucleotiodes can be produced in a short period of time with great accuracy

Question 15

How do bacteria reproduce?

Answer 15

Binary fission

Question 16

What does ‘in vivo’ mean?

Answer 16

in living organisms / cells

Question 17

What is a plasmid?

Answer 17

Small, circular DNA found in the cytoplasm of bacteria that is separate from the main DNA.

Question 18

Why are plasmids particularly useful in gene technology?

Answer 18

Can be transferred into bacteria easily. May contain genes for antibiotic resistance
that can be used as a genetic marker. Contains a replication origin so can replicate

Question 19

The same restriction enzyme is used to cut both the DNA fragment from the source
DNA and the DNA of the vector.
Why is this important?

Answer 19

Vector and fragment need to contain
complementary ‘sticky ends’ to allow them
to form a recombinant plasmid.