Gene Cloning Methods Comparison:

PCR:

-can amplify any piece of DNA without using cells like Sanger
-Heat is used to denature DNA
-Errors of PCR limits the number of good copies that can be made when large amount of gene are needed.
-DNA is incubated in a test tube with special DNA polymerase (Taq pol)
-DNA template, dNTP, primers (2 known sequence), and taq pol
-has repetitive steps of heating and then cooling DNA
-used in RFLP

Vector Cloning:
-Apply gene of interest into plasmid to bacterial cell
-DNA ligase glue cut sequence of DNA together
-Direct manipulation of genes for practical purposes
-gel electrophoresis and RFLP used to contain gene of interest 
-uses RE, vector DNA, and bacterial cells
-Takes longer than the other methods

Sanger DNA sequencing:
-can amplify any piece of DNA without using cells like PCR
-DNA template, dNTP, primers (1 known sequence), polymerase, and ddNTP
-the first letter of the new sequence is missing
-used to sequence DNA

All:
-all need DNA template 
-all produce new copies of DNA
-all manipulate DNA

DNA Translation

Initiation:

• For protein synthesis to begin, Initiation Factors assemble the small ribosomal sub-unit, mRNA, initiator tRNA, and large ribosomal sub-unit
• Small ribosomal sub-unit attaches to the mRNA near the start codon (AUG)
• Initiator tRNA (UAC) binds to the codon
• a large ribosomal sub-unit joins to form the active ribosome
• The 3 binding sites are A (Amino acid), P (Peptide), E (Exit)

Elongation:

• Protein synthesis occurs
• Polypeptide becomes longer
• when the initiator tRNA is bound to the ribosome, the A site is occupied by the tRNA
• Anti-codon base pairs with the second mRNA codon , ribosome can link the second amino acid together 
• Peptide bond forms between the first and second amino acids
• resulting dipeptide is attached to the tRNA at A site
• the mRNA moves along by one codon and enters the P site
• tRNA carrying the next amino acid enters the A site
• the polypeptide chain is transferred to the amino acids in the A site
• mRNA moves along by one codon and the tRNA which no longer carries an amino acid exits from the E site

Termination:

• the phase begins when a stop codon is reached on the mRNA
• the polypeptide and the components of translation are separated (KA-BOOM)
• A release factor (protein) cuts the polypeptide from the last tRNA
• the polypeptide will then fold into a 3D shape which is ready to perform its cellular activities