Download Extraction of RNA File

Lab. 13
Extraction of RNA
Ribo nucleic acid :This type of nucleic acid play abig role in transcription of genetic formation that
found in DNA and synthesis of protein.
The RNA will be different on DNA :
1)
The RNA molecule will be single strand.
2)
The RNA contain Ribose sugar.
3)
The RNA contain (U) Uracil instead of thymine , U lingage with A
4)
There are three type of RNA
5)
The RNA found in nucleus and cytoplasm but the DNA found in nucleus
only
6)
The RNA found virus .
The DNA will be double helix for that consist from two strands coiled with
together
There are three type of RNA
1)
Massenger RNA : it is function transport the information from DNA to
Ribosomes in sequances of amino acids in order to synthesis protein chain.
The synthesis process of mRNA from one chain of DNA as template called
transcription .
DNA
2)
3)
transcription
mRNA
Tranlation
Protein synthesis
Transfer RNA (t RNA) : this type found in cytoplasm and it is job to
transport the active amino acid to ribosomes and contact with mRNA .
There are number of tRNA more than amino acids number. Therefore
there are more than one tRNA for one amino acid.
Ribosomal RNA ( r RNA) : this type participate in building of ribosomes and
play role in uniting the amino acid to creating peptide chain.
Protein synthesis process:
4) In the beginning the first step represented by segregation the two
strands of DNA by some enzymes in order to transcription the
genetic material from DNA to RNA and used the one strand of DNA
as a template creating the m RNA strand by help of RNA polymerase
after thtat the mRNA will be leaving the nucleus to cytoplasm where
Ribosomes found.
5) The second step include activation or loading the amino acid by
some enzyme and contact the active amino acid with the tRNA that
special with it, after that the
6) t RNA transport to ribosomes and contact with m RNA the anti
codon that found on the tRNA.
7) The m RNA contain the nucleotides called codon and the codon
must be corresponding with anticodon , this way the amino acid will
be together in the ribosomes to great polypeptide by peptide bonds
after that the tRNA will be free to attached with another amino acid.
RNA Extraction by Trizol
Protocol:
1) Add I ml Trizol to the sample and homogenize
2) Add 200 µ incubate o chloroform to the homogenate
3) Vortex vigorously
4) Incubate on for 15 min.
5) Centrifuge to get phase saparation 12000g for 15 min. at 4 C
6) Transfer the agueous phase to a fresh tube
7) Precipitate the RNA by mixing with 0.5 ml isopropanol
8) Incubate on ice for 10 min.
9) Centrifuge for 10 min. at 12000 g at 4 C
10)
Remove the supernatant
11)
Centrifuge at 7500 g for 10 min. at 4 C
12)
Remove the supernatant
13)
Air dry
14)
Dissolve RNA pellet in appropriate volum of RNase –free H2O