Research Methodology &

Project Writing
(Life Sciences)

UGC BSR Faculty Fellow

 1
Many Definitions of Research

• Research in common parlance refers to a search for

knowledge.
• Research is a scientific inquiry aimed at learning new facts,
testing ideas, etc.
• It is the systematic collection, analysis and interpretation of
data to generate new knowledge and answer a certain
question or solve a problem.

In fact, Research is creativity/

new, innovative or novel idea!
 Types of Research

• Basic research vs Applied research

• Quantitative vs Qualitative
• Descriptive vs Analytical
• Conceptual vs Empirical (Experimental type)
• Primary Research vs Secondary Research

There are Some Other Types of Research

 Research Process in Flow Chart

Experiments

The research work over many years with doctoral students and early
career researchers is important and the work should be published in
the Internationally peer reviewed journals
Rules of the game of doing science – Research article or Project
 Preparation of Research Proposal for PhD

Project title
Research supervisor
Background (Introduction)
Objectives
Proposed methodology (Methods)
Expected research contribution (Outcome)
Work plan & Deliverables
Synopsis
Bibliography & Resources
Writing Research Proposal for PhD

 Writing a research proposal is both

science and art
 Writing a formal research proposal should
be started by the time one has decided on
the topic after searching literature for the
study and identified the gap
 A good research proposal is based on scientific facts
 Project title
Project Title is a name of the Project.

A proper project title describes the whole assignment

in one sentence.

The title of a project is of ultimate importance, thus

make sure to take your time to find the best one. 

Titles must be attractive and exciting at the same

time. The title must convey the meaning, the area of
intervention and the goals of the project while being
enticing.

Project titles makes you to understand the main goal

of the Project work and deliverables.
 Research Supervisor

Choosing a supervisor is tricky because you don't know

much about them until you start working with them

Know the academic background and research interests of

a scientist .
Explain why he is the perfect person to supervise you by
making reference to his work

Mention any funding you are applying for. Ask if you can
arrange a time to meet or talk on the phone about the
project.
 Background/Introduction
(Review of Literature)
Literature survey is needed to identify
a broad and sub-area of interest as well as
a specific topic of research
Background
The introduction serves the purpose of leading the reader from a general subject area
to a particular field of research.
Introduce your topic.
Write about the research you plan to carry out.
State your rationale.
Explain why your research is important.

Rational of the study- Hypothesis & Key questions

A research hypothesis can be defined as a clear, specific and predictive statement that
states the possible outcome of a scientific study.

Current status of research and development (International and National Status

(~ 2000 words)
Identifying a Broad Field of Research –
Basic and Applied

• Basic research – Plant Virology- Broad Area

Fundamental Research Topic, e.g. Isolation of a gene,
Understanding of apoptosis, Identification of a target gene in
a virus, a key gene that is involved in replication, etc.

• Applied research – Plant Biotechnology Broad Area

e.g. Development of genetically modified crop for insect
resistance (Bt cotton), a drug, vaccine, etc.
 Area of Interests
Choosing a research field, depending on your interest and
Academic background, You have a passion for a particular
subject and have noticed a gap in the current research?

In Life or Biological Sciences – there are many f areas

In different systems - Plant, Animal, Human, Microbial, etc.
1. Stem cell research
2. Clinical research
3. Diagnostics
4. Bioinformatics and Compulational Biology
5. Nanobiotechnology or Nanotechnology etc.
6. Cancer genetics
7. RNA interference
8. miRNA interference
9. Genome or gene editing etc.
Identification of Gap in an Area of Interest
or Problem Statement

A thorough search for literature to abreast with the latest

Developments in the field to avoid repetition of work
By using different key words, e.g., in RNAi in Plants – RNAi,
small RNAs, miRNAs, RNA silencing, RNAi constructs, Transgenic
RNA lines, Gene silencing, etc.

Various sites for literature search like Pub Med, Springer,

Elsevier, etc.

No access, but abstracts are freely available, which is sufficient

To know the developments in the field
 Narrowing down to a sub-area
(specific topic)
•Choosing a specific research topic, for example Deciphering the Signaling
pathway leading to Apoptosis in human cells, Identification of QTLs associated
with yield in rice, Targeting important genes for the control of colon cancer,
Functional validation of genes involved in fungal pathogenesis in Arabidopsis,
etc.

•Prioritizing the topic

 Development of working hypotheses
with key questions

• After extensive literature survey, researcher should find a

gap in the area of interest and state in clear terms the
working hypothesis or hypotheses/ questions.
• Working hypothesis is tentative assumption made in
order to draw out and test its logical or empirical
consequences.
• Express your research questions and hypotheses in a
focused clear manner
Examples of identifying gaps in an area of interest
RNAi discovery - hypothesis
In 1995 Guo and Kemphues
wanted to show that they had
cloned the C. elegans par-1 gene
(required for normal division of
the zygote). Used antisense RNA
par-1 to prove.

Antisense par-1 Sense par-1control

3’ 5’ 5’ 3’

Injection produced Injection produced

mutant par-1 phenotype mutant par-1 phenotype
What?
 In C. elegans (Guo & Kemphues, 1995)

 First noticed that sense RNA was as

effective as antisense RNA for
suppressing gene expression in worm C.
elegans.
 silencing effects are evident in both
injected animals and their progeny even
though RNA transcripts are rapidly
Degraded in the early embryo.

 Remained a mystery...
“The basis for the sense effect is under
investigation ...”.
Fire et al., Nature 391. (1998)
The discovery of
RNA interference

Working hypothesis: the silencing effect by antisense or sense RNA

might be due to low-level contaminations of double-stranded RNA.

Results:
• double-stranded RNA is far more effective than single-stranded RNA.
• The sense or antisense RNAs lose their silencing effect if they are purified from the
contaminating double-stranded RNA (dsRNA).
• Only a few molecules of dsRNA are required per cell ® non-stochiometric effect that implies a
amplification component.
 Research Objectives
• Based on the identified research gap, the specific objectives
have to be set out.
• This is a very important and pivotal section and everything else
in the study is centered around it
• The objective of the proposed study should be specific,
relevance, feasible and stated very clearly
• The objective stated should be specific, achievable and
measurable, avoidance of duplication

• Too many objectives to be avoided

• Even just one clearly stated relevant objective for a study would
be good enough
• If there is more than one objective the objectives can be
presented in the appropriate order of importance
 Defining the objectives
Aim - Control of Fusarium wilt in tomato by
silencing ornithine decarboxylase (ODC) gene of
the target fungal pathogen by RNA interference

1. Retrieval of ornithine decarboxylase gene (ODC) sequence

from NCBI, BLAST search to rule out any off-target effect,
Cloning and the preparation of hair-pin RNAi construct with
the selected off-target free target sequence
2. Generation of tomato transformants by Agrobacterium-
mediated genetic transformation using the developed RNAi
construct
4. Molecular analyses for demonstrating the transgene
integration and expression in RNAi transgenics
5. Testing of the promising RNAi transgenic T1/T2 lines for
resistance against Fusarium wilt caused by F. oxysporum
 Research Design (Methodology)
• Research or Experimental design is the process of planning a
study to meet specified objectives. Planning an
experiment systematically, properly and clearly is very
important in order to ensure that the right type of data and a
sufficient sample size and power are available to answer the
research questions of interest as clearly and efficiently as
possible.
• Research design is needed because it facilitates the smooth
sailing of the various research operations, thereby making
research as efficient as possible yielding maximal information
with minimal expenditure of effort, time and money.
 Proposed Methodology
(Methods -Techniques)
It is essential to discuss procedures clearly
and completely (in sufficient detail) with considerable
amount of details so that the methods can be reproduced

•Study or research design (appropriate methods)

•Replications - Biological (3) & Technical replications (3)
•Independent experiments (2 or 3)
•Positive and Negative Controls, Experimental group
•Principle of Local Control (local var as a check in breeding)
•Work place- site observation data collection (Lab experiments- infrastructure, e.g.,
Plant tissue culture lab, Field experiments )
•Instrumentation (make sure that the required equipment )
•Specific procedures or techniques (mol biol, cell cult, etc.)
 Proposed Methodology
(Methods -Techniques)

The Principle of Randomization (Randomized tables are

available)
Study population / Sampling specifications (race, region,
country, etc.)
Sampling design – size (~200 for medical research,
~2000 seeds for mutation studies)
Data collection methods
(questioner, experiments, primary & sec source etc)
Data analysis methods (Quantitative- Statical analysis –
Mean, SD, SE, t-test, Chi-square, Analysis of Variance and
Covariance, Regression, etc. using softwares &
Qualitative)
Data quality (reproducibility, honesty is important..)
 Proposed Methodology
(Methods -Techniques)
Alternative Strategies
(e.g. Besides the original experimental plan, one should also have an alternative
research approches)

Case Study Method

(e.g. a case study of a particular organisation or to describe a research method or
Biosafety
clinical work on a disease)
(IBSC & RCGM)
Ethical Consideration
(Ethics committee and Biosafety committee)
An example for experimental approaches
to address the set objectives

• Objective (Aim) - Control of Fusarium wilt in tomato by

silencing ornithine decarboxylase (ODC) gene of the target
fungal pathogen by RNA interference

1. Retrieval of ornithine decarboxylase gene (ODC) from NCBI

2. Subject the target ODC gene sequence for homology search by
BLAST to rule out any off-target effect
3. Select the off-target free region of the partial target gene
 NCBI sequence retrieval
In silico analysis for selection
Source of
sequence of partial target ODC gene
Homology search with” nr” data base
sequence of Fusarium
Gene
homology oxysporum
BLAST - 21bp fragments off-target search
Off-target
search
Prediction of putative siRNA from the selected
gene sequence through Target Finder
siRNA
prediction
m-fold software prediction - hairpin RNA
dsRNA
stability
check •Minimum homology,
•formed putative siRNA
Selected •Formed stable dsRNA in plant RNAi- FoODC
sequence
3. Cloning and the preparation of hair-pin RNAi
construct with the selected target sequence

Cloning of partial ODC gene from

F. oxysporum
560 bp
PCR product was confirmed by sequencing and it
was similar to F. solani

A XhoI XhoI EcoRI SwaI XhoI BamHI XbaI PstI

LB 35sP nptII PolyA 35sP Sense intron Antisense Ocs RB

ODC ODC PolyA
4. Generation of tomato transformants by
Agrobacterium-mediated genetic transformation
using the developed RNAi construct
 4. Molecular analyses for demonstrating the transgene
integration and expression in RNAi transgenics

Fungal ODC RNA lines expressed

the intended siRNAs
Kb M +C 10 16 21 32 56 57 60 61 67
1
0.5 M Wt 10 16 21 32 56
0.25 (a) 25 bp
21 bp
PCR screening 15 bp

M 60 61 67 69 72 76
(b) 25 bp
21 bp
15 bp

Southern analysis
4. Demonstration of the stability of the transgene in the
subsequent generations
5. Testing of the promising RNAi transgenic lines for resistance
against Fusarium wilt caused by F. oxysporum
6. Selection of the best transgenic event for open field trials

Control ODC-RNAi Line 89

 Expected research
contribution (Outcome)

The purpose of the  research

contribution statement is to summarize the
new contributions or expected results of the
proposal (in bullet form)
 Work plan &
Deliverables

Time frame (3 years) and work schedule

The proposal should include the sequence of

tasks to be performed, the anticipated length
of time required for its completion and the
Deliverables
 Synopsis

It gives a panoramic view of your

research for quick analysis by the
supervisor. 

It gives gist of your proposal

 Bibliography & Resources

A bibliography is an alphabetized list of all

the sources used in the project but not
too many!

Arrange bibliography alphabetically

Resources (Internet sources….)

Concept Project Proposal for Funding

Concept note (generally format is not given)

Title page, PI and contacts

Background information (Introduction)
Objectives
Experimental approach (Methods for each
objective)
Expected outcome
References
Brief bio-data of PI
Full-length Project Proposal for Funding
Full length project proposal
(Format is provided by the funding agency)

Title page, PI , Co-PIs and contacts

General information (Title, Institutions, Cost,
Coordinator)
Project summary
Particulars of investigators
Technical details
Introduction (Origin of the proposal, Rational of
the study- Hypothesis & Key questions)
Current status of research and development
(International
and National Status & References)
Full-length Project Proposal for Funding Cond.

Relevance and expected outcome of the study

Preliminary work so far in the proposed work
Specific objectives (with verifiable indicators for
each objective)
Time line (Tasks and duration)
Research Methodology
Budget particulars (Institute-wise – for each PI)
Existing facilities (Institute-wise – for each PI)
Declaration/Certification
Detail Biographical sketch of investigators
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