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Backbone
这是一篇关于Backbone的思维导图，主要内容有Pocess、Products、Sources、recombinant DNA。
编辑于2022-06-11 16:11:36- Recombin…
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Fermentation
Pocess
Other techniques
Sterelization
Decimal reduction time of a contaminant organism
The time required at a certain temperature to kill 90% of the organisms being studied
depends on two variables
The species
The temperature
The suspending medium
a. The methophile
b. The thermophile
The comparation
viruses<mesophiles<fungal/yeast<Clostridia<Endospores
Temperature profile
batch sterilisation
Continous sterilization
Steam injection
Hold
1-5min at 140
up/down
seconds
Plate exchange
Hold
10min at 120
up/down
6s
1. Pretreatment
1. Sorting
2. sieving
3. hydrolysis
4. medium formation
5. sterilisation
2. Bioconversion
contrast with chemical reactions
Bioconversion
lower
temperature
pressure
energy imput
aqueous solution
enzyme
specific
pure product
single isomeric form
Chemical reactions
organic solvent
can be racemic mixture
Concepts
Biomass
metabolites
primary
secondary
differences
essential for survival
accumulation in cell's life
distribution in species
Overporduction
altered allosteric site
specificity of the enzyme
Biotransformation
By free/immobilized enzymes
By whole cells
3. Downstream processing
1. cell disruption
2. concentration
centrifigation
filtration
flocculation
precipitation
3. purification
Chromatography
crystallization
4. drying and packaging
Downstream Processing
1. Cell disruption
for the metabolite contained in inclusion body
Non-mechanical
small scale
drying
freeze
vaccum
osmotic shock
temperature shock
chemolysis
autolysis
use bacteria own enzymes
chemicals
EDTA, triton X-100
enzymes
hydrolyses
b-glucanases
antibodies
peniciline
cycloserine
Mechanical
large scale
ultrasonic disrupters
homogenisers
pressure cells
bead mills
2. Clarification
remove cellular debris
Centrifugation
Tubular centrifuge
Disk-stack centrifuge
Filtraction
Batch filtration
plate filters
Continuous filtration
rotating drum vaccum filters
3. Concentration
Evaporation
Precipitation
Ultrafiltration
using membranes
4. normal Purification
purify bulk product
do not requires great purity
liquid extraction
for small molecules
organic solvent
for complex molecules
aqueous two phase
Crystalization
supersaturation method
cooling
evaporation
seperated by filtration/cetrofugation
5. ultra Purification
Chromatography
types
adsorption
affinity
gel filtration
high performance liquid
hydrophobic
metal chelate
defination
concentrate and purify the culture broth
cost
40%-90%
culture broth contain 95% water
Products
Derivatives of pyruvates
propionic acids
cheese
acetone-butanol
acetic acid
vinegar
Lactic acid
yoghurt
cheese
Ethanol
beer
wine
bread
Biomass(cell)
yeast
bread macking
food supplement (dried)
animal feeds
Macromolecules
SSP
recombinant bacteria's products
Insulin
human growth hormone
enzymes
amylase
protease
cellulase
polymers
polysaccharides
Xanthan gum
food industry
Good suspending and stabilizing agent for oil/water emulsions
such as salad dressings.
petroleum industry
component of drilling muds
Dextran
medical drips
glucose polymer
beads act as molecular sieves
cross linked
Alginic acids
colloidal
food stabaliser and thickener
Primary metabolites
fermented foods
Soy sauce
mushrooms
sauerkraut
yoghurt
cheeses
vinegar
baverages
wine
distilled beverages
beer
Brewing
wine
grape juice
Beer
malted grain
process of brewing
Malting
maximum production of useful enzymes
Chemical changes
acidification
Protein degredation
proteinase
degredate big sized proteins
peptidase
degredate medium sized proteins
Starch degredation
Alpha-amylase
Beta-amylase
Amylase
Protease
minimum enzymatic activities and plant growth
first soak and kiln drying
Mashing
use the generated enzymes to degredate the starch and proteins
use warm water
in mashing tank
the wort is prepared for fermentation
Wort seperation
seperate wort(liquid) from solid part
solid contribution
large amounts of protein, poorly modified starch, fatty material, silicates, and tannins
filtration
boil wort
Spoiled batches of beer occurred from bacterial contamination of the ferment, contamination could be prevented by heating wort to 55 degrees C before adding yeast
pasteurization
add hops
Add hops
Extract flavors (bitter acids) and aromatic compounds
Sterilizes hopped wort
fermentation
CO2 vent out through a tube...
conditioning
filtering
packaging
distilled beverages
distillation of fermented solusion
production of alcoholic beverages and alcohol fuel through chemical mixing processes of infusion, boiling and fermentation.
lager
organic acids
lactic acid
Lactic acid fermented food
types
homofermentative
1 glucose to 2 lactic acid and 2 ATP
heterofermentative
1 glucose to 1 lactic acid, 1 ethanol and 1 ATP
Food
diary product
butter milk
traditional
cultured
acidified
Yogurt
Kefir
meat product
vegetable
Flavouring agents
aminoacids
monosodium glutamate
secondary metabolites
Vitamins
B12
riboflavin
antibiotics
citric acid
Types
Lactic acid
glucose to lactic acid
Propionic acid
lactic acid to propionic acid
Ethanolic
glucose to ethanol and CO2
Heterolactic
glucose to lactic acid, ethanol, and CO2
Alkaline
Ammino acids to ammonia
Sources
carbohydrates
Sugar crops
Starches
Lignocellulose
recombinant DNA
recombinant protein
health
Gene therapy
Vaccines
food
transgenetic crops
transgenetic animals
Applications
Development of disease-resistant plants
Food crops that produce greater yields
“Golden rice" engineered to be more nutritious
Genetically engineered bacteria that can degrade environmental pollutants
Large scale production of human proteins by
genetically engineered bacteria (e.g. Insulin).
Development of Vaccines
Gene Therapy for genetic diseases
Development of disease resistant crops
Genetic engineering
recombinant DNA
cloning vectors
plasmids
bacterial phage
Aspects we need to engineer
replication
transcription
translation
regulation
clone
Clone – a molecule, cell, or organism that was produced from
another single entity
Common of vectors
Marker genes
ori
MCS
multiple cloning site
recognition sites for several restriction enzymes in which insert is cloned into.
promotor
The variation of vectors
host
Prokaryotic
E.coli
Bacillus
Eukaryotic
yeast
Cosmid
COS ends of phage λ DNA
Then cosmid is packaged into viral particles and used to infect E. coli cells at low copy number
Shuttle vectors
allow cloned DNA to be
moved between unrelated organisms. A shuttle
vector is a cloning vector that can stably
replicate in two different organisms
Expression vectors
have been developed for both prokaryotic and
eukaryotic hosts. This type of vector allow high
level protein expression in bacterial cells
because they have a prokaryotic promoter site
next to the MCS (multiple cloning sites).
Applications
recombinant bacteria
how can we synthesize our animal protein?
get the mRNA from the animal tissue
reverse transcribe it into cDNA
than we direct ligate the cDNA into the cloning vector
human proteins as drug
hormone
insulin
antibodies
diagnosis?
recombinant animal cells
2 methods
Retrovirus-mediated transgenics
Infecting mouse embryos with retroviruses before
the embryos are implanted
• Size of transgene (transferred genetic material) is
limited
pro-nuclear micro injection
• Introduces the transgene DNA at the earliest
possible stage of development of the zygote
• DNA is injected directly into nucleus of egg or sperm
engineered animals
research
disease model animals
nematodes
zebra fish
fruit fly
mice
GFP
increase quality and yield of food
organ donors
drug synthesizers
Gene therapy
Recombinant virus
Subunit vaccine
living vaccine
DNA vaccine
Recombinant plants
methods
breeding(hybridization)
Sexual cross between two lines and repeated
backcrossing between hybrid offspring and parent
protoplast fusion
Protoplast cell is a callus cell whose cell wall has
been dissolved by the enzyme cellulase
leaf fragment techniques
• Small discs are cut from leaf
• Cultured in a medium containing genetically
modified Agrobacter (Agrobacterium tumefaciens)
– A soil bacterium that infects plants
– Bacterium contains a plasmid, the TI plasmid,
that can be genetically modified
– DNA from the TI plasmid integrates with DNA
of the host cell
• Leaf discs are treated with plant hormones to
stimulate shoot and root development
gene guns
Chloroplast engineering
Antisense technology
Polyploid plants
Increased size
• Vaccines for plants
• Genetic pesticides
Bt genes
normally from bacteria
• Herbicide resistance
• Enhanced nutrition
Golden rice
provitamin-A (beta-carotene)