BIO 325 Study Guide - Midterm Guide: Dna Ligase, Nucleic Acid Hybridization, Intron
Gel Electrophoresis
• Gel electrophoresis – DNA, RNA, proteins and SDS are all based on negative charges and
sizes
• Two types of gels
o Agarose (10 bp differences)
o Acrylamide (1 bp differences)
• Gels are molecular sieves
• Gels have pores of a specific average size
• DNA a e isualized i a gel ith ethidiu roide
o Large amounts of each fragment – bands
o Small amounts of each fragment – smear
Separating (fractionating) DNA Fragments
• If digesting an entire genome, there are many different sized fragments that can see a
smear
• Cut out the portion of the gel that contains the appropriate sized fragments for cloning
Plasmids
• Plasmid – a ateria otai sall irular DNA alled plasmid, whih otai
extra information
• Bacteria normally contain plasmids that replicate autonomously from the chromosome
• Plasmid cloning vectors can replicate human DNA fragments
• Cloning a fragment of DNA
o Transform – bacteria with plasmid
o Idetify trasfored oloies y seletio
Transforming Bacteria
• Not all vectors contain insert
find more resources at oneclass.com
find more resources at oneclass.com
Document Summary
If digesting an entire genome, there are many different sized fragments that can see a smear: cut out the portion of the gel that contains the appropriate sized fragments for cloning. Transforming bacteria: not all vectors contain insert, how to eliminate cells containing empty vectors, lac z converts colorless x-gal into a blue pigment, the cloning site is in the lac z gene. Inserts will disrupt the lac z gene and the colonies will be colorless on x-gal. Hybridization: probe, denaturation, renaturation (hybridization, a short single-stranded piece of dna that is labeled and used in hybridization e(cid:454)peri(cid:373)e(cid:374)ts is (cid:272)alled a (cid:862)pro(cid:271)e(cid:863) Genomic library summary: break up the genome into fragments, clone all of the fragments. Polymerase chain reaction (pcr: procedure do not use dna ligase (cid:862)pcr(cid:863, revolutionized molecular biology or genetics, can start with a single copy of dna and in few hours have billions, forensics, dna testing, embryo testing.
