biotech Flashcards
isolation of plant cell
cellulase for breaking the cell wall
isolation of fungi
chitinase for removing chitin
isolation of bacterial cell
lysozyme for breaking peptidoglycan
found in saliva, tears
isolation of DNA from protein rna and lipids
protease RNAse lipase
isolation of DNA from garbage
put it in chilled ethanol, it will float
genetic scissors
restriction endonuclease
what gel is used in gel electrophoresis
agarose gel (red algae)
charge on DNA
negative
which dye is used in gel electrophoresis
ethidium bromide
what are the steps for gel electrophoresis
- put the fragments in a bed containing agarose gel
- keep a +vely charged electrode, i.e, anode on the opposite end
- the dna fragments being -vely charged will get attracted to the anode and rush towards it.
- the smaller fragments will move fast in comparison to the larger ones
- once these have settled, put ethidium bromide dye on it
- now place a uv lamp near the bed, the dna will be visible as orange fragments.
what is elution
the process of separating the gene of interest from the gel
1st restriction enzyme
hind II - haemophilus influenzae
what are sticky ends
overhanging stretches of nucleotides
form h2 bonds with their complimentary cut counterparts
stickiness facilitates the action of DNA ligase…..
what are recognition sites
special sites where the restriction endonuclease cut the dna
what are the features of plasmids
double stranded dna always circular extra chromosomal non essential self replicating piece of dna
2 types of vectors
expression vectors
cloning vectors
what are the properties of vector
origin of replication- decides the copy number
selective marker
cloning site
how to separate transformant gene from non transformant
selective marker
name an artificial plasmid
pbr322
what is insertional inactivation
inactivation of gene for synthesis of enzymes
bacteria that infects plants with TI gene
agrobacterium tumifaciens
disarmed vector
a vector that could earlier harm the host, but cannot do so now. as dangerous gene is destroyed
what do retrovirus do
convert rna to dna
list the bt crops
tomato potato corn cotton- bollworm rice
genes in cotton bollworm
cry I AC
cry II AB
genes in cornborer
cry I AB
methods of molecular diagnosis
pcr
elisa
what is genetic engineering
techniques to alter the chemistry of genetic material
what is cloning
the ability to multiply copies of antibiotic resistance gene in E.coli
EcoRI
Escherichia coli RY 13
what are exonucleases and endonucleases
exonucleases remove nucleotides from ends of the DNA whereas endonucleases make cuts at specific positions within the DNA
how does a restriction endonuclease function
- inspects the length of DNA sequence
- once it finds its specific recognition sequence it will bind to the DNA and cut each of the 2 strands of the double helix at specific points in their sugar phosphate backbones
- each restriction endonuclease recognises a specific palindromic nucleotide sequence
what is ORI
origin of replication
sequence from where replication starts
any piece of DNA when linked to the sequence can be made to replicate within the host cells
what is transformation
procedure through which a piece of DNA is introduced in a host bacterium
what is selectable marker
helps in identifying and eliminating non transformants and selectively permitting the growth of transformants
how is a bacterial cell made competitive to take up DNA
- treating it with specific concentration of a divalent cation such as calcium, which increases the efficiency with which DNA enters the bacterium through pores in its cell wall.
- rDNA can then be forced into such cells by incubating them w rDNA on ice, followed by placing them briefly at 42°C [heat shock] and then putting them back on ice.
what is microinjection
rDNA is directly injected into the nucleus of an animal cell
what is biolistic/gene gun
cells are bombarded with high velocity particles of gold/tungsten coated with DNA
what is reqd in PCR
template strand
DNA polymerase (taq)
nucleotides
primer
steps in pcr
denaturation- the 2 template strands separate at 95°C
annealing- primers attach on both the strands when the temperature is lowered to 55°-65°C
extension- when the temp is raised to 72°C new strands are formed
what are generically modified organisms
plants animals bacteria or fungi whose genes have been altered by manipulation are called genetically modified organisms
what is gene therapy
collection of methods that allows correction of a gene defect that has been diagnosed in a child
what are transgenic animals
animals that have had their DNA manipulated to possess and express an extra gene
how do we benefit from transgenic animals
normal physiology and development study of disease biological products vaccine safety chemical safety testing
