Molecular Genetic Analysis and Biotechnology Flashcards
Characteristics of restriction enzymes
180 axis of symmetry
Recognize a 4 nt sequence or a 6 nt sequence
Gene Cloning (3 steps)
Digest chromosmal and vector DNA with the same restriction enzyme
Mix. Sticky ends will form base pairs (anneal)
Seal the nicks with DNA ligase
2 Types of vectors
Cloning vectors and shuttle vectors
Standard cloning plasmid
small, circular, site of origin, antibiotic resistance gene
Expression Plasmid
specialized vector can overproduce the protein encoded by the gene
Shuttle vectors
contain origins of replication for two organisms
DNA library
test tube containing fragments of every gene in an organism
DNA probe
radioactive DNA fragment that is complimentary to the gene you want to clone
Southern blot
Probing for DNA fragment using a DNA probe that is complimentary to the gene of interest
Northern blot
Probing for RNA fragment using a DNA probe that is complementary to the gene (transcript) of interest
Western bloth
probing for protein using antibodies that are specific for the protein of interest
PCR
used to amplify specific regions of DNA.
RFLP Mapping
can be used to identify genetic defect
DNA Sequencing
used to determine nucleotide sequence of any gene
can show gene mutation
Dideoxy Sequencing
ddNTPs are used to cause chain termination
four seperate reactions