Lecture 11: Biotechnology 2 Flashcards
Biotechnology can improve the human condition through
- treatment of disease
- industrialisation/ resource efficiency e.g. of food processes, clothing & washing industry
- caring for our environment
- improving nutrition
Biotchnology can use:
-prokaryotic cells
-single-celled eukaryotes (e.g. yeast)
cell cultures from higher eukaryotes (e.g. Hela cells, plant protoplasts cultures)
-Whole organisms (e.g. transgenic plants & animals)
example of biotechnology for treatment of disease
diabetes -INSULIN
Insulin:
- first protein to be sequenced
- normally produced in pancreatic beta cells
- Removes excess glucose from the blood
- Injections of insulin can control type 1 diabetes (and also type ll in some cases)
- used to be isolated from bovine and swine pancreas (cattle & pig)
synthetic insulin from
the 1960’s
-became the first approved genetically engineered pharmaceutical product in 1982
__ ways insulin can be made
two
-expression in E.coli or yeast
Method 1 for forming insulin: Chains A & B are produced from two plasmids in two bacterial strains
- Insulin chain A & B
- take purified chains form both (purified chain A and purified chain B)
- place together = disulphide bonds form
method 2 for forming insulin : Proinsulin is produced in yeast
proinsulin : A B + C (C joins A + B)
- proinsulin is made and disulfide bonds form
- a protease removes the 33 amino acid chain C, purification
DNA sequencing wasn’t invented until the
1970’s
PCR wasn’t invented till
1983
human genome wasn’t sequenced till
2003
Eukaryotic gene can be too ___ to amplify using PCR
LONG
PCR can be used for genes up to about 10 kb
-human genes can range from 200bp - 2,500,000 bp in length
genome fragmentation by
restriction enzymes
fragments packages into
Bacterial Arificial Chromosomes of 100-300 kb
-amplified bacterial lineages derived from a single bacterium with a single plasmid can be stored
how to find your gene:
- Nucleic acid hybridisation can be used to find your gene of interest in a BAC library
- works on the principle of hybridisation of complementary DNA sequences
- radioactive or luminescent probe to identify
Industrialisation/ resource efficiency e.g. of food processes, clothing & washing industry how does it help & e.g.
used because it streamlines a process and makes it cheaper and / or more reliable
-Rennet
-Cheese production requires the use of
proteases
how was proteases traditionally isolated?
as a complex of enzymes from veal calf stomachs
1g of extract can coagulate
15litres of milk. Equates to 0.3mg of enzymes per kg cheese
how is rennet now sourced?
- chymosin B gene transferred to bacteria & fungi e.g. fungi Aspergilus niger & Kluyveromyces lactis
- Protein produced by fermentation then extracted
Advantages to new source of rennet
higher yield
better texture
less bitter
Social considerations to new source of rennet:
vegetarian
halla
kosher
(but not organic)
plant biotechnology looks at
- increasing productivity
- increasing storage of crop
- increasing nutritional or taste attributes
- disease resistance
- nitrogen fixation for crops
- stress resistance
methodology - plant transformation: Naked DNA can be
‘shot’ into cells but vectored DNA is often used to make stable transformations
methodology - plant transformation:
Agrobacterium :
transfers part of a plasmid (T-DNA) into plant cells
methodology - plant transformation:
Agrobacterium tumefaciens
- plasmid produces plant hormones to induce callus tissue that excretes sugars that the bacteria uses to grow
- disarmed so as not to produce crown galls
- T-DNA can then be used to transfer genes of interest into plant
methodology - plant transformation:
Agrobacterium rhizogenes :
-similar infection strategy to A. tumefaciens
- leaf discs or rott sections infected
- hairy roots form
- can induce shoot formation with plant hormones cytokinin and auxin
improving nutrition biotech example:
golden rice
Golden Rice:
- vitamin D deficiency is a major problem in developing countries
- normal rice ontains no Beta-carotene (provitamin A)
to synthesise Beta-carotene in rice grains what needs to be added
2 enzyme
- Phytoene synthase
- Carotene desaturase I
phytoene synthase:
- Promoter gt1 (endosperm specific)
- coding region from daffodil
Carotene desaturase l:
- promoter gt1 (endosperm specific)
- Coding region from bacterium Pantoea ananatis
Golden rice stil needs to be equivalent to normal rice in:
- yield
- disease resistance
- stress resistance
- quality and storage
- available in local varieties (done through introgression)
- insertion in a neutral locus
biotechnology can utilise both
prokaryotic and eukaryotic cells and organisms
