recombinant DNA applications

Question 1

what is rDNA?

Answer 1

recombinant DNA
- a DNA constructed in a lab from 2+ natural or artificial DNA fragments
- fragments are ligated using a DNA ligase

Question 2

how does molecular cloning work?

Answer 2

• DNA fragment to be cloned is inserted into a vector by ligation, forming a vector-insert-complex
• antibiotic resistance gene also added
  -> only bacteria containing plasmid with chosen DNA will survive
• ## complex is introduced into host cells of a bacterial colony

Question 3

what does the term transformation mean in the context of recombinant DNA technology?

Answer 3

introduction of the DNA complex into host cells
-> transfection if host cells are from an animal

Question 4

what is the word for a host cell altered by transformation?

Answer 4

transgenic

Question 5

what are 3 uses of DNA cloning?

Answer 5

• biopharmaceuticals
  -> used to make human proteins w biomedical application, eg insulin
• gene therapy
  -> functional form of a particular gene provided to cells of a patient’s body who usually has non-functional gene
• gene analysis
  -> us DNA cloning to help understand how genes in an organism function

Question 6

what are cDNA libraries?

Answer 6

complementary DNA libraries
- collections of bacterial clones containing cDNA representing expressed genes in a given tissue at specific conditions and corresponding to a specific physiological situation

Question 7

what are gDNA libraries?

Answer 7

genomic DNA libraries
- collections of bacterial clones containing fragments of genomic DNA digested by a restriction enzyme

Question 8

how are cDNA libraries constructed?

Answer 8

1. RNA extraction and mRNA purification
2. cDNA synthesis -> production of double stranded DNA
3. adapters are ligated and digested
4. ligation of cDNA into the vector
5. packaging of rDNA (cDNA in vector) into phage capsule
6. plating of phage library int host bacteria

Question 9

how are gDNA libraries constructed?

Answer 9

1. gDNA extraction n purification
2. digestion of gDNA and vector w a restriction enzyme
3. ligation of DNA fragments into digested vector
4. packaging of rDNA (gDNA fragments inserted into vector) into phage capsule
5. plating phage library into host bacteria

Question 10

what are probes?

Answer 10

labelled RNA or DNA fragments w a known sequence used to detect a complementary sequence in DNA or RNA population

Question 11

where does a homologous probe come from?

Answer 11

from the same organism

Question 12

where does a heterologous probe come from?

Answer 12

from a different organism, used to detect similar sequences in the studied organism