Lahore University of Management Sciences

BIO 300 Methods on Molecular Biology

Instructor
Room No.
Office Hours
Email
Telephone
Secretary/TA
TA Office Hours
Course URL (if any)

Fall 2013

Dr. Muhammad Tariq, Dr. Saima Anwar, Dr. Zakir Ullah & Dr. Aziz Mithani
9-415 A , 9-420A , 9-413 A , 9-419 A
N/A
m.tariq@lums.edu.pk, Saima.anwar@lums.edu.pk ; zakirullah@lums.edu.pk; aziz.mithani@lums.edu.pk
8218, 8352, 8217, 8397
N/A
N/A
N/A

Course Basics
Credit Hours
Lecture(s)
Recitation (per week)
Lab (if any ) per week
Tutorial (per week)

Course Distribution
Core
Elective
Open for Student Category
Closed for Student Category

4
Nbr of Lec(s) Per Week
Nbr of Rec (s) Per
Week
Nbr of Session(s) Per
Week
Nbr of Tut(s) Per
Week

N/A
N/A

Duration
Duration

N/A
N/A

Duration

4 hours/day

N/A

Duration

N/A

Yes
No
Biology Majors
-

COURSE DESCRIPTION
This lab based course introduces modern tools of molecular cloning; protein expression and purification which are used to understand
cell and molecular biology of cellular function. In addition, students are introduced to experimental model system Drosophila
melanogatser which is used to perform classical genetics by crossing various genetic mutants as well as different tissues of fruit fly
larvae are dissected to understand development, inducible gene expression, active and silent genes. Different modules of the lab
course are designed so that students address a specific question by utilizing different recombinant DNA technology tools. Students will
design and carry out experiments to address specific questions about genes involved in the regulation of transcriptional regulation,
cellular development, cell fate determination and cell death etc.

COURSE PREREQUISITE(S)

BIO100, BIO101
BIO216, BIO221, BIO212
Instructors consent

COURSE OBJECTIVES

Introduce students to modern concepts and techniques in molecular cloning and protein purification
Introduce students to form a scientific question and address it by different tools possible
Introduce genetics, molecular, biochemical and cytological approaches to address same scientific question

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Learning Outcomes
Students must be able to:
Develop a scientific approach to address a biological question about cellular function

Perform experiments independently by having a specific protocol and must also be able to search for experimental protocols

Address a cellular function through multiple approaches like genetic, molecular and biochemical.

Grading Breakup and Policy

There will be no Midterm and Final Exam. The whole grading will be based on lab and will be decided by individual instructor for his/her modules
Assignment(s):
Home Work:
Quiz(s):
Class Participation:
Attendance:
Midterm Examination:
Project:
Final Examination:

Examination Detail

Midterm
Exam

Yes/No: NO
Combine Separate:
Duration:
Preferred Date:
Exam Specifications:

Final Exam

Yes/No: NO
Combine Separate:
Duration:
Exam Specifications:

COURSE OVERVIEW
Week/
Lecture/
Module

Topics
1.

Module 1

Module 2

2.

Drosophila as a Model for Genetics and

Development
Drosophila Embryonic Staging.
Dissection of IIIrd Instar Larvae for isolation
of Imaginal Discs.
Different stages of Drosophila development.
Balancer Chromosomes and markers
Inheritance pattern of Eye Color in
Drosophila

3.

Chromosome Mapping of a Transgene

(Hsp83EGFP)

1.

Polytene Chromosomes: Identification of

heat shocked Puffs and Banding Patterns

2.

Developmental Tissue Specific Gene

Expression

Recommended
Readings
Selected readings for each
module from books
mentioned in
Textbook(s)/Supplementary
Readings

Objectives/
Application
Students will be introduced to basic
genetic tools which can be used to
answer different questions by using
Drosophila melanogaster as a model
organism

To study the temporal and tissue

specific gene expression in vivo.
Students will also learn what are

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3.

Identification of Heterochromatin and

Euchromatin regions on Polytene
Chromosomes

Module 3

Induction of Somatic Clones using Flp-FRT system:

Effect of Homozygous Mutations of Polycomb on
Expression of Homeotic Genes (Ubx).

Module 4

Chromatin Immunoprecipetation (ChIP).

Module 5

Reverse Genetics: To see the effect of knockdown of

Polycomb and Trithorax group proteins on their target
genes in cell culture

Module 6

Module 7

Module 8

Module 9

Introduction
Preparation of chemically competent E. coli
cells
Bacterial Transformation with plasmid DNA
Plasmid purification
Gel electrophoresis agarose
Restriction analysis and plasmid mapping
Plasmid preparation (restriction and
dephosphorylation)
Purification of vector DNA from agarose gel
Ligation
Transformation and plating
Identification of recombinant clones
(inoculate O/N cultures)
DNA isolation from buccal swabs and
chicken, cow and sheep liver
Amplification of mitochondrial DNA cox-2
gene segment via PCR
Identification of recombinant plasmids
Mini-preparation of plasmid DNA
Digestion with restriction enzyme
Purification of PCR products for DNA
sequencing
Multiplex PCR of dystrophin gene
Analysis of multiplex PCR products on
polyacrylamide gels
Transformation of E. coli for protein
expression Isolation of total
RNA from HeLa cells
RT-PCR
Grow E. coli for protein expression (collect
cells)
Affinity purification of recombinant protein
Separate fractions on SDS-PAGE
Coomassie stain half; electroblotting of the
other
Western blotting
Expression and purification of E. coli RNA
polymerase

epigenetic marks which can be used

to
differentiate
between
euchromatin and heterochromatin
regions of chromosomes.
Students will learn how they can
study the phenotype of recessive
lethal mutations
Students will learn a technique
which can be used to study the
binding profiles of transcription
factors or any other DNA binding
proteins
They will learn how dsRNA can be
used to silent the target genes at
post-transcriptional level (PTGS) in
Drosophila cell culture.

Basic Molecular biology tools kit:

They will learn how they can clone
and express their gene of interest.

Basic Molecular biology tools kit:

Students will learn basic protein

biochemistry

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Module 10

Isolation and characterization of satellite

DNA from bovine genomic DNA

Textbook(s)/Supplementary Readings

A Handbook of Drosophila Development (1982) Robert Ransom (ed.). Elsevier Biomedical

Ashbumer, M. (1989) Drosophila: A Laboratory Handbook and Drosophila: A Laboratory Manual Cold Spring Harbor Laboratory
Press
Drosophila. A Practical Approach (1998) Roberts, D.B. (ed.). IRL Press
Lawrence, P. A. (1992)The Making of a Fly: The Genetics of Animal Design Blackwell Scientific Publications
Rubin, G. M. (1988) Drosophila melanogaster as a ExperimentalOrganism. Science 240, 1453-1459
Ralph J. Greenspan (1997) Fly Pushing CSHL Press
Campos-Ortega, J. A. and Hartenstein, V. (1997) The Embryonic Development of Drosophila melanogaster. Springer-Verlag
(Berlin, Heidelberg).