CH. 7 DNA STRUCTURE AND REPLICATION

IIR v IIIS features (ie. Which is virulent or outcome for mice injected with it)

IIR didn't kill, IIIS kills, heat killed IIIS didn't kill, heat killed IIIS and IIR kills, meaning IIR transformed into IIIS.

Know the assembly of polynucleotide chains.

Individual nucleotides are assembled into chains by DNA polymerase, phosphodiester bond forms between 3' hydroxyl group of one nucleotide and 5' phosphate of adjacent, each polynucleotide chain has a sugar-phosphate backbone made of alternating sugar and phosphate groups.

Be able to identify the characterization and functions of the 5 types of DNA Polymerase and what properties they have.

Pol I - more in cell than II or III, removes primer and fills gaps Pol II - repairs DNA damage caused by external forces and at replication forks Pol III - largest (900 kD), in vivo 5'-3' DNA replication proofreading Pol IV and V - repair DNA damages caused by external forces (UV, mutagens, radiation)

What are the similarities and main differences in DNA replication between prokaryotes and eukaryotes?

Similarities dsDNA unwound at replication origins, replication forks are formed, bidirectional synthesis creates landing and lagging strands. Differences More DNA in euk than prok Chromosomes in euk are linear Euk DNA is complexed with proteins

What is the function of the sliding clamp?

Sliding clamps can close around dsDNA during replication to enhance the processivity of polymerases. It anchors DNA pol III enzyme to the template.

What is the difference between complementary and antiparallel? Be able to identify if given sequences. How are they important for DNA functions?

The bases are complementary. Why important: allows DNA replication, gene expression, meiotic pairing, and molecular hybridization. The strands run antiparallel. Why important: allows DNA replication, transcription and repair.

What causes the gap in the telomeres and what enzyme fixes this problem? Know basics of telomerase activity steps.

The lagging strand has a gap preceding the Okazaki fragment with 3'-OH. Polymerase filled the gap.

The _____ IIR cells were _____ into _____ IIIS cells.

rough; transformed; smooth

Be able to identify a replication fork, the 5'-3' orientation of the original DNA strands; show RNA primers, DNA polymerase, Okazaki fragments, and directions of replication on lagging and leading strands.

see review

Cancer cells maintain _____ activity and are _____.

telomerase; immortal

Know the steps of DNA replication in their correct order and know the basics for each step.

1 DNA molecule undergoes replication I → 2 DNA molecules, undergoes mitosis I → 2 separated DNA molecules, undergo replication II → 2 DNA molecules each

Transformation is used by _____ to transfer DNA.

bacteria

Differences between dNMPs and dNTPS.

dNMPs - deoxynucleotide monophosphates part of the polynucleotide chain have single phosphates dNTPs - deoxynucleotide triphosphates NOT part of the polynucleotide chain

_____ are not part of a polynucleotide chain.

dNTPs

Avery, MacLeod-McCarty used extracted heat-killed SIII bacteria that was divided into aliquots and treated to destroy either DNA, RNA, proteins, or lipids and polysaccharides

just mem

The transforming factor carried hereditary information but could not identify the molecule which led to Avery-MacLeod-McCarty in vitro experiments.

just mem

Know the proteins and genes associated with those we went over in class.

primase - dnaG gyrase - gyrA/B RNA pol - rpoB DNA pol I/II - polA/B DNA helicase - rep DNA ligase - lig single-stranded binding protein - ssb

What are Chargaff's rules and how do they correlate to the base composition studies critical conclusions?

A=T, C=G; purines = pyrimidines

Define base stacking, major and minor grooves, an identify the 3 forms of DNA and their properties.

Base stacking - offsetting of adjacent base pairs so their planes are parallel. Major grooves (12 Angstroms) and minor grooves (6 Angstroms) alternate and are regions where DNA binding proteins make contact with nucleotides. 3 Forms of DNA: B-DNA: Watson-Crick model, low-salt conditions, right-handed A-DNA: mainly in vitro, high-salt conditions, right-handed Z-DNA: found near transcription start sites, zigzag, no major groove, left-handed

When did transformation not occur and this led to what discovery?

DNase treated IIIS filtrate. Transformation still occurs with RNase and Protease treated.

Why do prokaryotes only have one origin of replication, while eukaryotes have several of them?

Euk DNA is much larger so it needs more origins of replication to finish in a timely manner.

Can eukaryotic organisms contain multiple ori on chromosomes? Be able to identify replication bubbles and forks from electron micrograph.

Eukaryotic chromosomes have multiple origins of replication.

What are the basics of DNA replication and what are the three attributes shared by all organisms?

Basics: semiconservative, bi-directional, polymerase III does main synthesis, direction is 5'-3', starts at point(s) of origin 3 attributes: Each strand of the parental DNA molecule remains intact during replication Each parental strand serves as a template for formation of an antiparallel, complementary daughter strand Completion of replication results in 2 identical daughter duplexes, each with one parental and one daughter strand

What is a consensus sequence?

Consensus sequences are nucleotide sequences most often found at each position of DNA in the conserved region.

Name the three theoretical models of DNA replication

Conservative: one daughter duplex has both parental strands and the other daughter duplex has both daughter strands Semiconservative: each daughter duplex has one parental and one daughter strand Dispersive: each daughter duplex has mixed parental and daughter strands

Know the 4 different subunits of DNA Pol III and what function is.

Core enzyme - polymerization activity Gamma complex - loads enzyme onto the template at the replication fork (clamp loader), requires ATP Beta subunit - prevents core enzyme from falling off the template Tau subunit - dimerizes two core polymerases to facilitate simultaneous synthesis on both strands

What is the chemical structure differences between RNA and DNA? (Hint: 2'-C)

DNA has thymine RNA has uracil

Explain the Meselson-Stahl experiments and how they ruled out the conservative and dispersive replication models in bacteria.

Meselson-Stahl experiment ruled out conservative replication 15N is heavy, 14N is light Process to determine semiconservative: Gen 0: E. coli DNA labeled with 15N - centrifuge shows all 15N/15N (heavy) Gen 1: Grown in 14N medium, cells replicate once - centrifuge shows all 15N/14N (hybrid) Gen 2: cells replicate twice - centrifuge shows half 14N/14N (light), half 15N/14N (hybrid) Gen 3: cells replicate third - centrifuge shows 3:1 ratio of 14N/14N(light) : 15N/14N(hybrid)

What are the 3 properties the genetic code has to have?

Must replicate Must encode info Must be able to change/mutate

What are the differences between Nucleosides and nucleotides? Be able to identify if given the name of one as an example and if it is a nucleoside or nucleotide. (Hint the ending will help you)

Nucleoside - nitrogenous base + pentose sugar How to identify: ends in -sine Nucleotide - nucleoside + phosphate group How to identify: all end in acid

What are Okazaki fragments and how are they combined after being synthesized? Know enzyme involved and what strand they are located on.

Okazaki fragments are formed on the lagging strand by RNA primers and are joined together by DNA ligase

Regarding DNA replication in prokaryotes, explain the following terms: Origin of replication Replication fork Circular chromosome Replication bubble

Origin of replication - where DNA replication proceeds Replication fork - where helicase splits DNA strands Circular chromosome - only found in prokaryotes, no telomeres Replication bubble - expansion around origin of replication between two replication forks

PCR amplification and dye terminator sequencing similarities and differences.

PCR amplification and dye terminator similarities: - Both have supply of four standard dNTPs PCR amplification and dye terminator differences: - PCR uses dsDNA, dye term uses ssDNA

What is RNA priming?

RNA priming is a universal feature of initiation of DNA replication in both euk and prok done by RNA polymerase, which synthesizes a short RNA primer complementary to the DNA template, primase does not need free 3' end

Name the main characteristics of telomeric sequences. How are telomeres important in chromosomes?

They are at the ends of euk chromosomes, don't contain genes, and contain special repeats (TTAGGG). They function by capping chromosomes and preserving their integrity and stability, protecting chromosomes from end-to-end fusions, protects from exonucleases and degradation, and prevents gradual loss associated with RNA-primed replication.

Why does proofreading need to occur? What activity allows DNA Polymerase to work?

To fix replication errors, which leads to DNA mismatches and inability of mismatched bases to form appropriate H bonds; 3'-5' exonuclease activity

List the 3 main points of the Transforming Principle.

Transforming agent is DNA Transformation is a genetic event, meaning changes affect the next generations Broad range of variation, meaning it can involve many different bacteria and traits

What is the most significant nucleoside phosphate?

Triphosphates: Precursor molecules for nucleic acid synthesis in the cell Adenosine (ATP) and guanosine triphosphates (GTP) are essential for energy

What causes and controls supercoiling?

Unwinding of chromosomes during DNA replication causes supercoiling; DNA Topoisomerases unwinds