Biol 335 midterm summary
Recombinant DNA technologies:
To study genes and genetic material
To engineer organisms for study
For mass production of useful biological molecules
Gene therapy
Terminology:
▪ Cloning: a collection of methods that allow dna to be inserted into
vectors that can be grown in bacteria. The cloned DNA can be
manipulated for stud or production.
▪
▪ Vectors: a DNA molecule/vehicle that is used to deliver foreign
genetic material into host cells where can be replicated and/or
expressed. (plasmids, bacteriophages, viruses)
▪ Cell cultures: to grow larger amounts of DNA and to express gene
products
▪ Restriction endonucleases and ligases: cut and paste DNA
fragments together
▪ Polymerase chain reaction: to amplify sequences of interest or act
as a diagnostic for the presence or absence of sequences
,▪ Site-directed mutagenesis: to study the impact of a sequence on
gene function
▪ CRISPR/CAS9: for gene editing and the study of gene function
Restriction endonucleases:
• Bacterial endonucleases recognize and cut at short palindromic
sequences DNA sequences. This generated sticky or blunt ends.
,• Bacterial DNA is typically methylated to protect its own sequences
from its own restriction endonucleases.
Ligases:
Enzyme that facilitates the joining of DNA strands together by
catalyzing the formation of a phosphodiester bond.
DNA sequencing
Sanger: chain terminating inhibitors
Gilbert: use chemicals that cleave preferentially at each base
Vectors:
Bacterial plasmids:
• Small circular extrachromosomal elements
• Replicate independently of the bacterial chromosome
, • Carry genes for bacterial resistance
• Easily exchanged between bacteria
• Many copies expressed per bacterial cell
Viral vectors
• Enter cells, replicate and express their genes as part of their
natural infection cycle.
• Can be engineered to improve safety and express cloned genes.
• Viral sequences may still be manipulated in plasmid form.
Features of a plasmid:
Features that would be useful:
Application Goals Feature
Recombinant DNA technologies:
To study genes and genetic material
To engineer organisms for study
For mass production of useful biological molecules
Gene therapy
Terminology:
▪ Cloning: a collection of methods that allow dna to be inserted into
vectors that can be grown in bacteria. The cloned DNA can be
manipulated for stud or production.
▪
▪ Vectors: a DNA molecule/vehicle that is used to deliver foreign
genetic material into host cells where can be replicated and/or
expressed. (plasmids, bacteriophages, viruses)
▪ Cell cultures: to grow larger amounts of DNA and to express gene
products
▪ Restriction endonucleases and ligases: cut and paste DNA
fragments together
▪ Polymerase chain reaction: to amplify sequences of interest or act
as a diagnostic for the presence or absence of sequences
,▪ Site-directed mutagenesis: to study the impact of a sequence on
gene function
▪ CRISPR/CAS9: for gene editing and the study of gene function
Restriction endonucleases:
• Bacterial endonucleases recognize and cut at short palindromic
sequences DNA sequences. This generated sticky or blunt ends.
,• Bacterial DNA is typically methylated to protect its own sequences
from its own restriction endonucleases.
Ligases:
Enzyme that facilitates the joining of DNA strands together by
catalyzing the formation of a phosphodiester bond.
DNA sequencing
Sanger: chain terminating inhibitors
Gilbert: use chemicals that cleave preferentially at each base
Vectors:
Bacterial plasmids:
• Small circular extrachromosomal elements
• Replicate independently of the bacterial chromosome
, • Carry genes for bacterial resistance
• Easily exchanged between bacteria
• Many copies expressed per bacterial cell
Viral vectors
• Enter cells, replicate and express their genes as part of their
natural infection cycle.
• Can be engineered to improve safety and express cloned genes.
• Viral sequences may still be manipulated in plasmid form.
Features of a plasmid:
Features that would be useful:
Application Goals Feature
