LAB #4
Part 1: Extractions
&
Part 2: Karyotyping

• From “chromo” and “soma” meaning  colored-body
• Chromosomes carry genes
• Found inside nucleus of human and Drosophila cells

FUN FACT ABOUT DNA!!!
 DNA is super coiled .
 DNA of chromosomes in one human somatic cell would be
6 feet long if stretched out!!!
 Human Genome = about 3 billion base pairs http://www.youtube.com/watch?v=4PKjF7OumYo

Human vs . Drosophila Chromosomes
Note: Polytene chromosomes of Drosophila .
Maternal & paternal pairs are closely oriented & all 8 chromosomes are joined at their centromeres in a region called the chromocenter.

Human & Drosophila Chromosomes
What do they look like under a microscope?
• You will view both under the DEMO Microscopes located on the side bench.
• Record sketches in your lab notebook.

Human / Fly Comparison
Humans
Flies
Drosophila &
Sarcophaga
Total
# of Pairs
#
Autosomal
Pairs
23 22
# Sex
Pairs
1
Total # of
Chromosomes
46
4 3 1 8

Part 1: Chromosome Extractions!
Today:
• Extract chromosomes from the SALIVARY GLAND of
Sarcophaga “Blowfly” larva .
• We will also use fruit flies larva.
• Instructor will demonstrate procedure = micro-dissection requires patience & care !
• You will be graded on effort and end result for this portion of the lab  Prepare 2 + chromosome squashes.

3 rd instar
LARVA

Sarcophaga larva – closer look http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm

Sarcophaga larva
Appearance on a dissecting microscope.
• Locate the mouthparts & salivary gland position.
Photo credit: David B. Fankhauser, Ph.D.
Professor of Biology and Chemistry
University of Cincinnati Clermont College. http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm

Using the dissecting microscope, “tease” away the fatty tissue, organs (e.g. trachea), etc. from the salivary gland.
Photo credit: David B. Fankhauser, Ph.D.
Professor of Biology and Chemistry
University of Cincinnati Clermont College.
Can you locate the salivary gland among the other tissues in this picture?
Hint: look for the translucent structure that contains “cells”! It may be bi-lobed, unless you cut it already.

Sarcophaga - Salivary Gland
Photo credit: David B. Fankhauser, Ph.D.
Professor of Biology and Chemistry
University of Cincinnati Clermont College. http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm

Sarcophaga –
Salivary Gland
( up close, dissecting scope)
Photo credit: David B. Fankhauser, Ph.D.
Professor of Biology and Chemistry
University of Cincinnati Clermont College. http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm

Sarcophaga –
Salivary Gland Isolated & Stained
Photo credit: David B. Fankhauser, Ph.D.
Professor of Biology and Chemistry
University of Cincinnati Clermont College. http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm

Sarcophaga –
Salivary Gland Squashes
Almost!
Not squashed enough!
Photo credit: David B. Fankhauser, Ph.D.
Professor of Biology and Chemistry
University of Cincinnati Clermont College. http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm

Sarcophaga –
Salivary Gland Squashes
Perfect…your goal for today!
Photo credit: David B. Fankhauser, Ph.D.
Professor of Biology and Chemistry
University of Cincinnati Clermont College.
You must use a compound light microscope for viewing these chromosome squashes (on slides!).
http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm

Karyotyping from a METAPHASE SPREAD
Chromosomes arrested in metaphase stage of mitosis.

A KARYOTYPE
• Definition: pictorial representation of chromosomes organized in pairs and sorted by:
1. Size
2. Banding Pattern
3. Centromere location

Karyotyping – Ideotype
Arranged according to:
1) Size,
2) Banding Pattern,
3) Centromere location

Chromosomes & Centromere Location

Human chromosomes are divided into 7 groups + sex chromosomes:
• Group A: 1-3
Large metacentric 1,2 or submetacentric
• Group B: 4,5
Large submetacentric, all similar
• Group C: 6-12, X
Medium sized, submetacentric
• Group D: 13-15
Medium-sized acrocentric
• Group E: 16-18
Short metacentric 16 or submetacentric 17,18
• Group F: 19-20
Short metacentrics
• Group G: 21,22,Y
Short acrocentrics

 Banding patterns generated by
Giemsa & trypsin stains.
 Giemsa stains chromosomes in regions of high [A] & [T].

(You must know these by name, specifically).

Phenotypically, male or female ?
Trisomy 21 or Down Syndrome : note the extra copy of Chr. #21.

Phenotypically, male or female ?
Klinefelter Syndrome (XXY) : note the extra sex chromosome.

Phenotypically, male or female ?
Turner Syndrome (XO) : note the missing sex chromosome.

Phenotypically, male or female ?
Jacob’s Syndrome : note the extra sex chromosome (Y).

Mini Fun Fact!
Polyploidy in humans most likely leads to miscarriages. 
Polyploidy in plants occurs quite frequently and the plants are completely functional 
(30 – 70% of plants today!)
Polyploidy: note extra copies of each chromosome.
Triplicates of each instead of pairs.

Let’s Play…..

• Phenotypically female
• Characterized by:
• Broad neck
• Webbed necked
• Broader shoulders
• Swelling of hands and feet
Turner Syndrome (XO)

• Phenotypically normal
• Characterized by:
• Dough like skin
• Tongue that appears swollen
• Mental Retardation
Down Syndrome
(trisomy Chromosome 21)

• Not seen in living humans
• Miscarried; Still birth
• Common in plants
Polyploidy
(multiple chromosomal copies)

• Phenotypically Male
• Appear normal
• Usually only found through genetic testing
• Arthur Capshaw = The
Genesee River Killer
• Gave this syndrome a bad rap
Jacob’s Syndrome (XYY)

• Phenotypically Male
• Characterized by:
• Reduced fertility
• Lanky youth stage
• Some possible breast tissue development mainly later in life
Klinefelter Syndrome
(XXY)

You can try these at home for extra help!
http://gslc.genetics.utah.edu/units/disorders/karyotype/ http://www.biology.arizona.edu/human_bio/activities/karyotyping/karyotyping.html
http://www.DNAi.org

Today’s Agenda:
Part 1: Chromosome Extractions (worth up to 10 points)
 Work in pairs. Complete before starting Part 2.
 While one salivary gland is staining, begin preparing
1 or more additional extractions (just in case the first doesn’t work out).
 Upon completion, have instructor “sign-off” in your lab notebook. Grade range 0 – 10.
 You will be graded on your best attempt – both end result & effort count 

Today’s Agenda, continued:
Part 2: Karyotyping (worth up to 10 points)
 Work in pairs.
 Practice karyotyping using magnetic boards.
 Then, complete (2) paper karyotypes:
1. One normal: male or female .
2. One abnormal:
• Trisomy 21, Klinefelter’s, or Turners (TA will assign).
 Observe DEMO slides: fly vs. human chromosomes
 Complete lab notebook tables & sketches.
 If time is limited, you may submit completed paper karyotypes next week.