Biotechnology
Raj Mutharasan
Dept of Chemical Engineering
Drexel University
Products of Biotechnology
Structure of Bacteria
Live cell pics at : http://www.cellsalive.com/
Structure of an Animal Cell
Source: http://www.biosci.uga.edu/almanac/bio_103/notes/may_15.html.
Biology Basics
DNA makes RNA makes Protein
• Nucleic acids (deoxyribonucleic acid (DNA) and ribonucleic acid
(RNA)
• DNA: Adenine and Guanine (both purines) and Cytosine and Thymine
(both pyrimidines)
• RNA: Adenine and Guanine (both purines) and Cytosine and Uracil
(both pyrimidines)
Biology Basics
Pairing of Nucleotides
A single strand of DNA
DNA makes RNA makes Protein
nucleotides are
linked by 3’ to 5’
phosphodiester
bonds
phosphate group on 5’ of
sugar linked to 3’
hydroxyl group of the
preceding sugar
Double Stranded DNA
Amino Acids – Back bone of Proteins
Genetic Code
Example
DNA
ATG
RNA
UAC
AA
Tyr
Transcription and Translation
An Overview
Restriction Enzymes
Over 3000 restriction endonucleases are known. The most useful for cloning work are
known as Type II which generally make a staggered cut within the recognition site and
across the double stranded DNA. The enzymes are named by a three letter code
indicating the organism from which they were isolated. For example, EcoRI was
isolated from E. coli and BamHI from Bacillus amyloliquefaciens.
The action of EcoRI is as follows:
5’- - G AATT C - - 3’ - -C TTAA G - - 5’ - -G- 3’O H
3’ - -CTTAA-
AATTC - 3'OH-G - -
Enzyme recognizes a six-base pair sequence in the DNA, and the cleavage results in singlestranded ends (sticky ends) that would readily reform hydrogen bonded base pairs again
with each other. Restriction endonucleases recognize four, five or six base pair sequences.
Examples: Restriction Enzymes
Process Overview
St ock Cu lt u r e >>
Sh a k e Fla s k s
St e r ilize
Fe r m e n t or &
M e d iu m
Se e d
Fe r m e n t or
P r od u ct ion
Fe r m e n t or
Ce ll
Se p a r a t ion
P r od u ct
P u r ifica t ion
Stoichiometry of Growth
What are cells made of?
Element
C
O
N
H
P
S
K
Na
Ca
Mg
Cl
Fe
others
% DW
50
20
14
8
3
1
1
1
0.5
0.5
0.5
0.2
0.3
A Good Approximation –
“Formula for Cell”
CH1.8O0.5 N 0.2
Concept : Cell Yield
Experimental observation –
Cell mass is proportional
to available substrate
300
250
Slope = 7.2 µg/ml per mM
200
150
100
50
0
0
10
20
Glucose, mM
30
40
Concept : Growth Reaction
Growth Can be represented by:
Considering primary constituents:
In general:
Cell Yield – Formal Definition
Cell Yield is:
Yx / s
Change in Cell Mass

Substrate Consumed
Yx / s
dX

dS
Similarly, product yield is defined as:
YP / S
dP

dS
Kinetics of Growth
Cell Growth in Batch Culture
Cell Growth - Kinetics
During log phase, growth kinetics is expressed as:
X  X0 e
t
ln( 2)
Doubling Time  t d 

Typical Growth Rates
Organism
E. coli
Yeast
Hybridoma
Insect Cells
Growth
Rate
µ [h-1]
2
0.3
0.05
0.06
Doubling
time
[h]
0.35
2.3
13.9
11.6
Metabolic Quotients
1 g S + YO2/S g of O 2 + YNH3/S g of NH 3 = YX/S g o
YNH3/S g of NH 3 = YX/S g of Biomass + Y
CO2/S
rs
rO 2
rx


 1 Yx / s  YO 2 / s
CO2 + others
Metabolic Quotients - II
General Definition:
Organism
qglucose
qO2
g/(g h) g/(g h)
E. coli
2.5
0.3
Yeast
0.5
0.2
Hybridoma
0.2
0.02
What is your “food” metabolic quotient?
Biotechnology – The next steps
This module gives a basic introduction to principles of
biotechnology and biochemical engineering.
With bit more background in biology, you can pursue
further reading in gene therapy, human genomics,
tissue engineering, cancer diagnostic tools, use of
biotechnology in crime detection, and a whole score of
other applications.