Biology 1107 - Lab Practical 2

Purpose of a lab report

To communicate research

7 sections of a lab report

Title, abstract, introduction (with hypothesis), materials and methods, results, discussion, references

Peer review functions as a form of ____

Quality control

Describe the peer review process

Scientist submits manuscript. Editors send the paper to peer reviewers. Reviewers give feedback on if the paper qualifies for publication. Author revises and resubmits.

Peer review ensures that

Only high quality studies that follow guidelines are published

Monomer of a protein

Amino acids

Basic structure of amino acid

Amino group, carboxyl group, r group

During polymerization of a peptide, what type of reaction occurs?

Condensation reaction

What bond forms between two amino acids?

Peptide bond

When amino acids have a peptide bond, they form...

An amide group (O-C_N bonded in the center)

Biuret's reagent contains what element?

Copper (II) sulfate

What reacts with the copper in the biuret's regeant?

The amide groups of proteins

At what pH do copper and amide groups react?

at a basic pH

What color does biuret reagent turn?

Violet blue color

De novo

From scratch

Why did we use biuret assay?

It's expensive and easy to prepare/use

Con of biuret assay

It's not as sensitive as other methods

Biuret assay will only detect polypeptides with ___ or more amino acids

3

A spectrophotometer measures..

The amount of light absorbed by a substance

Biuret Assay absorbs light best at...

550 nm

Proteins are ubiquitous, meaning..

They're found inside the cell

Detergents are amphipathic, meaning..

They have hydrophobic and hydrophilic parts

What happens when a detergent comes in contact with a membrane?

Displaces the phospholipid membrane

Detergents form _____ in aqueous solutions

Micelles

Variables on the assay standard curve

Known concentration and optical absorbance

The standard curve in the assay lab was used to determine

Protein concentration of an unknown substance

Detergent used in experiment

SDS - sodium dodecyl sulfate

BSA stands for

bovine serum albumin

Gene

Sequence of DNA that determines a trait

Alleles

Inherited variations of a gene

Genotype

An individual's combination of alleles

Phenotype

The physical presentation of a trait

Gene located on autsomes

Autosomal Inheritance

Sex linked inheritance

Gene is located on the X chromosome

Species of fly

Drosophila melanogaster (written with underline)

Why use fruit flies?

small, short life cycle, easy to maintain generations, produce many offspring, many distinguishable traits

Life cycle of a fruit fly

egg, larva, pupa, adult

Male flies have

Sex combs

Two major parts of cell cycle

Interphase and mitotic phase

Subphases of interphase

G1, S, G2

What occurs in s-phase?

DNA synthesis (creation of sister chromatids)

Phases of mitosis

PPMAT

Phrophase

Chromosomes condense (dark circle)

Prometaphase

Nuclear envelope breaks down (see chromosomes spread)

Metaphase

Chromosomes line up

Anaphase

Chromosomes separate

Telophase

chromosomes condense in opposite ends

Cytokinesis

Cleavage furrow/ cell plate forms, cell splits

Structures of mitosis to remember

nucleus, nucleolus, chromosomes, spindle apparatus, cell plate, asters, spindle fibers, equator, cleavage furrow

An allele that occurs normally

Wild-type

An allele that occurs abnormally

Mutant

A dissecting microscope is used to

View larger objects

Total magnification of dissecting microscope

10x

Father of modern genetics

Gregor mendel

Species name for garden pea

Pisum sativum

Who first studied genetics using fruit flies?

T.H. Morgan

Fruit fly wild type eye

Red

Fruit fly mutant eye

White

Male flies are ____ than females

smaller and have darker tips

Female abdomen tips are..

More pointy, lighter, striped

Life cycle of fly is

10-12 days

Flies hatch in...

24-36 hrs

Eye trait in flies was

sex-linked recessive

Zygote

single fertilized cell

P generation of flies

White eye females (rr) x red eye males (R)

F1 generation of flies

Red eye females (Rr) and white eye males (r)

White eyes are...

Sex-linked recessive

F2 generation

Females: (Rr and rr) Males: (R and r)

DNA Fingerpinting

Technique for identifying individuals based on their unique DNA

Polymorphism

Two or more variant forms of base pairs that cause different phenotypes among individuals

Steps in DNA fingerprinting

1. Isolate genomic DNA, 2. Amplify polymorphic regions, 3. Cut DNA into smaller fragments, 4. Visualize DNA fragments and determine sizes

DNA can be obtained from..

Any tissue or biological fluid

Polymerase Chain Reaction (PCR)

Research technique that produces millions of copies of the same segment of DNA

First step of PCR cycle

Denature: DNA strands separated by heat

Second step of PCR cycle

Annealing: DNA primers bind to single strands

3nd step in PCR cycle

Extension: DNA polymerase synthesizes new strand using free dNTPs, nucleotide monomers

Rate of DNA synthesis is..

Exponential

Bacteria can protect themselves from viruses by

Creating restriction endonucleases - recognize and cut specific base pair sequences

We can use restriction endonucleases to..

Cut replicated polymorphisms, create fragments of varying lengths

Sticky ends

when cut, there is an end of unpaired bases that are "sticky"

What did we use in the experiment to visualize DNA fragments?

Agarose gel electrophoresis

Agarose is a..

Polysaccharide - dissolves in water and gels as it cools

What is the comb used for in the gel?

Creates wells in the liquid agarose that dries solid

DNA sample is loaded with a..

Loading buffer

An _______ ________ is applied across the gel

Electric current

DNA is _____ charged

Negatively

Because of DNA's charge, it moves _____ the black electrode and _______ the red electrode

Away from the black electrode (negative), towards the red electrode (positive)

Smaller DNA fragments move ______; Larger ones move ______

Faster; slower

What is used to visualize the DNA fragments?

Ethidium bromide, a mutant - Inserts between the bases of double stranded DNA

Ethidium bromide is viewed under a ______

UV light - and it's very sensitive

DNA marker

1 kb (kilobase) "ladder", used to count known lengths of DNA fragments

Markers on the DNA marker are measured in

bp (base pairs)

Well defined "line" of DNA

Band

DNA lane

Corridor for which gel leaves a well

Developer and year of PCR

Kary Mullis 1983

Monomer of DNA

deoxyribonucleoside triphosphates

Denaturing DNA separates the _____

Double strand into single strands

Elongation

actual synthesis of DNA strands

EcoR1 cuts

GAATTC

Sal1 cuts

GTCGAC