Ans 1. Some of the important recombinant proteins and their therapeutic uses are listed below:
|Recombinant proteins||Therapeutic uses|
2. DNase I
3. Tissue Plasminogen activator (t-PA)
4. Anti-thrombin III
6. Interferon alpha (INF alpha)
7. Interferon Beta (INF Beta)
8. Interferon gamma (INF gamma)
9. Factor VIII
10. Factor IX
|1. Diabetes mellitus
2. Treatment of cystic fibrosis
3. Acute myocardial infection
4. Prevention of blood clots formation in heart patients
5. Reversal of acute kidney transplantation rejection
6. Used for hepatitis C (as vaccine)
7. Multiple sclerosis
8. Chronic granulomatous disease
9. Treatment of Haemophilia A
10. Treatment of Haemophilia B
Ans 2. Name of the restriction enzyme: B am H1
The substrate DNA on which it acts:
The products it produces:
Ans 6. Shake flasks are used for growing and mixing the desired materials on a small scale in the laboratory. A large scale production of desired biotechnological product is done by using ‘bioreactors’. Besides better aeration and missing properties, the bioreactors have following advantages:
- Small volumes of cultures are periodically withdrawn from the reactor for sampling.
- It has a foam control system.
- It has become and pH control systems.
Ans 7. Palindromic nucleotide sequences in the DNA molecule are groups of bases that from the same sequence when read both forward and backward. Five examples of palindromic DNA sequences are as follows:
- 5’______G G A T C C ______3’
5’______C C T A G G ______3’
- 5’______A A G C T T ______3’
5’______T T C G A A ______3’
- 5’______A C G C G T ______3’
5’______T G C G C A ______3’
- 5’______A C T A G T ______3’
5’______T G A T C A ______3’
- 5’______A G G C C T ______3’
5’______T C C G G A ______3’
Q 10. Describe briefly the followings:
- origin of replication
- downstream processing.
Ans 10. a. one of the major components of a plasmid is a sequence of bases where replication starts. It is called origin of replication (ori). This is a specific portion genome that serves as start signal for self-replication (to make another copy of itself). Under narutal conditions, each plasmid replicates to produce 20-30 copies per cell. This number can be artificially increased. Any piece of DNA when linked to this sequence can be made to replicate within the host cells. This property is used to make a number of copies of linked DNA (or DNA insert)/ It is desirable that cloning of target DNA should be carried out in such a vector where origin facilitates high copy number.
- Bioreactors are large vessels (having a volumes of 100 to 1000 liters) which are used for biological conversion of raw materials into specific products individual enzymes etc., using microbial plant, annual or human cells. Each bioreactor has a cylindrical stirred tank to facilitate the mixing of contents. The stirrer provides facility of mixing the contents as well as the availability of oxygen. A bioreactor is more advantageous than shake flasks. It has an agitator system to mix the contents properly, an oxygen delivery system to make availability of oxygen, a foam control system, a temperature control system, a pH control system and a sampling port to withdraw the small volumes of the culture periodically.
- once the foreign gene product is synthesized, it is collected from the bioreactor and then processed further for its separation and purification. The purified and finally finished product is now ready for marketing. After biosynthetic stage, the collection of product, its separation and purification is known as downstream processing. Before marketing, the product has to be formulated with suitable preservatives. In case of drugs obtained from recombinant proteins, clinical trials and strict quality control testing must be carried. These quality control testing and clinical trials vary from product to product.
Q 11. Explain briefly
- Restriction enzymes and DNA
Ans 11. A. Polymerase chain reaction (PCR) is a technique of synthesizing multiple copies of the desired gene (or DNA) in vitro. This technique was developed by Kary mullis in 1985. It is based on the principle that a DNA molecule, when subjected to high temperature, splits into two strands due to denaturation. These single stranded molecules are then converted to original double stranded molecules. By synthesizing new strands in presence of enzyme DNA polymerase. Thus, a double stranded molecule of DNA is duplicated and multiple copies of the original DNA sequence can be generated by repeating the process several times.
The basic requirements of PCR are –
Two nucleotide primers.
- Restriction enzymes, which cleave DNA duplex at specific points in such a way that single-stranded free ends project from each fragment of DNA duplex. These single-stranded free ends are called sticky ends because they can join similar complementary ends of DNA fragment from some other source.
- Chitinase. It is it is an enzyme used to dissolve fungal cell wall. It results in the release of DNA along with several other macromolecules and impurities.
Q 12. Discuss with your teacher and find out how to distinguish between
a. Plasmid DNA and Chromosomal DNA
b. RNA and DNA
a. Plasmid DNA is a piece of symbiotic DNA present in addition to chromosomal DNA mostly in bacteria and yeast. It does not from the part of normal chromosomal DNA of the cell. Plasmid DNA is capable of replicating independently.
b. Differences between DNA and RNA
|1. It is mainly confined to the nucleus. A small quantity occurs in mitochondria and chloroplasts.
2. Its quantity is constant is each cell of a species.
3. It contains deoxyribose sugar.
4. Its pyrimidines are adenine and thymine.
5. The amount of adenine is equal to the amount of thymine, also the amount of cytosine is equal to the amount of guanine.
6. It consists of 2 polynucleotide chains held together by hydrogen bonds, and coiled into a double helix. Some viruses () have single-stranded DNA.
7. Its molecular weight varies from 2 to 6 million.
8. It is of 2 types: linear intranuclear and circular extranuclear.
9. It can replicate itself.
10. It controls structure, metabolism, heredity, differentiation and evolution.
11. It is a component of chromosomes.
12. It is a genetic material in all organisms.
13. It does not contain unusual bases.
14. A primer is needed for replication.
15. Its renaturation for replication.
16. It transfers its information to mRNA (transcription).
17. DNA is hydrolyzed by the enzyme DNA-ase.
|1. It mainly occurs in the cytoplasm. A small quantity is found in the nucleus.
2. Its quantity varies in different cells.
3. It contains ribose sugar.
4. Its pyrimidines are adenine and uracil.
5. Adenine and uracil are not necessarily in equal amounts, nor are cytosine and guanine necessarily in equal amounts.
6. It consists of a single polynucleotide chain. It may fold on itself and get hydrogen-bonded and coiled into a pseudohelix. Some viruses (rheovirus) have double-stranded RNA.
7. Its molecular weight varies from 25,000 to 2 million.
8. It is of 3 types: mRNA, tRNA, rRNA. Each type is further of many subtypes.
9. It can’t replicate itself. It is formed by DNA, some RNA viruses (paramyxo virus) can produce RNA from RNA template.
10. It brings about protein synthesis. It also starts replication.
11. It is a component of ribosomes.
12. It is a genetic material in certain viruses.
13. It may contain unusual bases in addition to the normal ones.
14. No primer is needed for transcription.
15. Its renaturation after melting is quick.
16. mRNA transfers its information to polypeptide (translation).
17. RNA is hydrolyzed by the enzyme RNA-ase.