BBMB 405 1st Edition Lecture 27 Outline of Last Lecture XIV Chapter 28 DNA Replication Repair and Recombination D Many types of DNA damage can be repaired Outline of Current Lecture XIV Chapter 28 DNA Replication Repair and Recombination E DNA recombination plays important roles in replication repair and other processes XV Chapter 29 RNA Synthesis and Processing A Review B RNA Polymerases catalyze transcription Current Lecture XIV Chapter 28 DNA replication repair and Recombination E DNA recombination plays important roles in replication repair and other processes 1 Recombination a Creates genetic diversity b Exchange of genetic information between two different pieces of DNA Share same gene with each other Reciprical recombination both receive DNA from the other c Essential in these processes Restart stalled replication process Repair double stranded breaks Provides diversity by exchange of DNA during meiosis Generates diversity in antibodies Integrates genetic material of some viruses into host s DNA Use for generation of knockout mice These notes represent a detailed interpretation of the professor s lecture GradeBuddy is best used as a supplement to your own notes not as a substitute d 2 Strand invasion RecA 3 4 5 6 a b Synapse DNA is brought close to each other c RecA Forms nucleoprotein filaments opens origin of replication attaches to pink blue and purple strands Double strand break repair occurs because of error in replication a Homologous recombination HR use sister chromosome to repair break Accurate repair can introduce specific sequence changes present in homologous DNA Recognize recession of 5 end on both sides then one undergoes strand invasion repair then serve as template for other strand Occurs at same time as replication but not during G1 b Non homologous end joining NHEJ backup used in G1 Error prone leads to indel random insertion or deletion Has no template so error prone DNA repair mechanism that can add or delete base pairs Genetic Editing a Engineer genome by double strand breaks Find gene of interest and target Results in indels Need a frameshift of anything but 3 Can also introduce separate DNA that contains mutation b Process Site specific endonuclease designed to cleave gene of interest Double strand break introduced by endonuclease cleavage NHEJ or HR repair leads to mutation NHEJ creates random mutations indels HR makes specific mutation and requires introduction of homologous DNA CRISPR Cas9 able to site specific sequences and target certain genes programmable RNA guided site specific endonuclease Holiday Junctions a Crerecombinase circularizes genome covalent 3 phosphotyrosine intermediate b 7 NExt XV Chapter 29 RNA Synthesis and Processing A Review 1 RNA and roles in gene expression a Replication priming RNA telomerase RNA b Transcription riboswitch long noncoding RNA c Translation ribosomal RNA transfer RNA signal recognition particle RNA transfermessenger RNA d Gene regulation small interfering RNA micro RNA e Processing small nuclear RNA small nucleolar RNA ribonuclease P self splicing introns f Genome integrity Piwi interacting RNA retrotransposons g Other RNA genomes CRISPR RNA circular RNA 2 RNA are processed post transcription Splicing chemical modification addition trimming cleavage editing 3 RNA transcription is catalyzed by RNA polymerase hydrolysis of NTP provides energy for creation of phosphodiester bond formation B RNA polymerases catalyze transcription 1 Hydrolysis of NTP provides energy for creation of phosphodiester bond formation 2 Initiation transcription bubble unwinds 17 base pairs and maintains by reanealing base pairs already transcribed 3 Elongation fidelity not as high don t need accuracy level of DNA once fall off it can t restart polymerase needs to get through entire sequence 4 Subunits of RNAP and function in bacteria eukaryotes are more complex as always a b alpha beta beta prime and omega are core subunits in that they stay part of the complex through out all of transcription c Alpha RNAP assembly interaction with promoter DNA interactions with regulatory factors d Beta and beta prime active site interactions with DNA RNA e Omega stabilizes assembled RNAP f Sigma transcription initiation part of RNAP holoenzyme but dissociates during elongation 5 RNA polymerase uses two divalent metal ions similar to DNA polymerase in that ther is active nucleotide and stabilizes leaving group 6 DNA is unwound locally at transcription bubble a DNA template bases exposed within bubble b DNA duplex re anneals as template moves out of bubble c Nascent RNA is separate from template d Elongation occurs at 50 nt s
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