Exopolysaccharide producing rhizobacteria and their role in moisture stress alleviation. Mr ...

113 Pages·2013·2.45 MB·English

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Exopolysaccharide producing rhizobacteria
and
their role in moisture stress alleviation.

Mr. Devendra Singh

Division of Microbiology
Indian Agricultural Research Institute
New Delhi – 110012, India
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Exopolysaccharide producing rhizobacteria and
their role in moisture stress alleviation.
By
Mr. Devendra Singh
A Thesis Submitted to the Faculty of Post-Graduate School,
Indian Agricultural Research Institute, New Delhi
in partial fulfillment of
requirements for the degree of
MASTER OF SCIENCE
IN
MICROBIOLOGY
2013

Approved by:
Chairman : Dr. Geeta Singh _______________

Co-chairman : Dr. Annapurna k. _______________

Member : Dr. Aruna Tyagi _______________

: Dr. Pramila Aggarwal _______________
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Division of Microbiology
Indian Agricultural Research Institute
New Delhi – 110012, India
Dr. Geeta Singh
(Principal Scientist)
CERTIFICATE
This is to certify that the thesis entitled
Exopolysaccharide producing Rhizobacteria and their
role in moisture stress alleviation. Submitted to the Faculty
of the Post-Graduate School, Indian Agricultural Research
Institute, New Delhi, in partial fulfillment of the
requirements for the award of MASTER OF SCIENCE
IN MICROBIOLOGY, embodies the results of bona fide
research work carried out by Mr. Devendra Singh Roll No:
20185 under my guidance and supervision and that no part
of this thesis has been submitted for any other degree or
diploma.
The assistance and help availed during the course of
investigation as well as source of information have been duly
acknowledged by him.
Date:
Place: New Delhi (Dr. Geeta Singh)
Chairman
Advisory Committee
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Dedicated to my parents
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Acknowledgements
I am beginning in the name of God who reincarnated me every morning with
strength, courage and confidence. I unassumingly bow my head before Lord
Almighty for his blessings and grace showered on me during my life and the
course of this study. I wish to record my sincere appreciation and gratitude to
Dr. Geeta Singh, principal Scientist, Division of Microbiology, New Delhi
and the Chairman of the Advisory Committee, for providing me an opportunity
to work under her guidance and suggesting me a challenging topic for the
research work. I am thankful to her for her valuable guidance, constructive
criticism and wholehearted support and encouragement to me throughout the
period of my study.
It is a great occasion for me to express my deep sense of gratitude
towards Dr. Annapurna K., principal Scientist, Division of Microbiology and Co-
chairperson of my Advisory Committee for her scholarly guidance, experienced
words, disciplined and constant encouragement led me to complete this venture. I
wish to extend my profound gratitude to member of the advisory committee,
Dr. Pramila Aggarwal, Division of Agricultural Physics, IARI, New Delhi, Dr.
Aruna Tyagi, Principal Scientist, Division of Biochemistry, IARI, New Delhi for
her tremendous guidance and enthusiasm shown towards my research work. I
extend my sincere gratitude to Dr. A. K. Saxena, Head, Dr. Dolly Wattal Dhar,
Professor, Division of Microbiology, for providing me the necessary
facilities and encouragement throughout this study It is my pleasure to express
my gratitude and reverence to Dr. Lata, Dr. Sangeeta Paul, Dr. Radha
Prasanna, Dr. Livleen Shukla, Dr. G. Abraham, Dr. Sunil Pabbi, Dr.
Surender Singh, Dr. Ramakrishnan Bala , Dr. Swarnalaxmi, Dr. Archna
Suman, Dr. Sunita Gaind and all other scientists of Division of Microbiology for
their encouragement and support throughout this endeavor. I extend my sincere
gratitude to Dr. Supradeep saha, Department of Agricultural Chemical, IARI ,
New Delhi, for providing me the necessary facilities and encouragement
during my research work. The acknowledgement is incomplete if I don’t say
thanks to all the staff of the Division of Microbiology and CCUBGA, especially
Rana sir, Kapoor madam, Surendra, Ravi, Gulati madam, and all others for all
the help they willingly rendered.
I am grateful to my senior Dashrath sagar, Samadhan sir, Bandeppa sir,
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Sarita mam , Sangeeta mam, Arjun sir, Borase sir, Ajinath sir, Vikas sir,
Anil sir, Krishnashis Das sir, Bala sir, Ramanand sir, Momocha sir, Dolamani
sir, Vasna sir, K. Reddy sir, for their great company in making my days of M.Sc.
in the division memorable. I am especially grateful to my classmates Jairam,
Dharmendra, Y.P.Reddy. I am also especially grateful to PRSA group and
Candid thanks to all my friends and juniors. The endless love, affection,
sacrifice and constant inspiration from my father, Mr. Tulsi Ram and my
mother, Mrs. Resham have enabled me to reach the footstep of my long
cherished aspiration. I owe a lot to them Thanks to all my family members for
their love and support. Finally, the financial assistance provided by the
I.C.A.R. in the form of I.C.A.R. Fellowship during the tenure is gratefully
acknowledged.

New Delhi

30-06-2013 Mr. Devendra Singh
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CONTENTS

Chapter Title Page No.

1 Introduction 1-6
2 Background 7-20
3 Materials and Methods 21-33
4 Research paper I 34-47
5 Research paper II 48-56
6 Discussion 57-61
7 Summary and Conclusion 62-63
Abstract (English) i-ii
Abstract (Hindi) i
Bibliography I-IX
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LIST OF TABLES

After
Table
Particulars page
No. No.

Research Paper- I

1 Quantification of EPS produced by different isolates of Wheat 40
crop rhizosphere under hydric stress condition (45% PEG )
2 Quantification of EPS produced by different isolates of Mung 40
crop rhizosphere under hydric stress condition (45% PEG )
3 Quantification of EPS produced by different isolates of maize crop 40
rhizosphere under hydric stress condition (45% PEG )
4 Morphological and Biochemical test of the selected isolates
41
Effect of different carbon source on EPS production by G7
5 (Pseudomonas sp)
42
Effect of different concentration of sucrose on EPS production by
G7 (Pseudomonas sp)
6 42
Effect of different C:N on EPS production by G7 (Pseudomonas sp)
7 42

8 Effect of different pH on EPS production by G7 (Pseudomonas sp)
43
Estimation of carbohydrate and protein content of the EPS
9 43
Analysis of FT-IR absorption spectrum of EPS Produced by
10 Pseudomonas sp. 43
Analysis of FT-IR absorption spectrum of EPS produced by
11 Microbacterium resistens 43

Analysis of FT-IR absorption spectrum of EPS produced by
12 Bacillus cereus 44
Research Paper-2

Effect of EPS positive bacterial inoculants on RAS/RT in wheat
13 rhizosphere at different soil moisture levels at 20 DAS 51
9

14 Effect of EPS positive bacterial inoculants on RAS/RT at 52
different moisture levels at 40 DAS
15 Effect of EPS positive bacterial inoculants on MWD at different 53

moisture levels at 20 DAS
16 Effect of EPS positive bacterial inoculants on MWD at different 53

moisture levels at 40 DAS
17 Effect of EPS positive bacterial inoculants on WSA at different 54

moisture levels at 20 DAS
18 Effect of EPS positive bacterial inoculants on WSA at different 54

moisture levels at 40 DAS
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LIST OF FIGURES

Figure Title After page
No.
No.
Research Paper-1

Comparison of EPS production ability under normal 40
and hydric stress condition
1

16S rDNA amplification from the selected bacterial 41
2
isolates

Phylogenetic tree constructed based on 16S rDNA 41
3
sequences of isolates

4 Dendrogram representing genetic similarity of bacterial 41
community of soils obtained from 16S rDNA PCR
5 FT-IR absorption spectrum of EPS produced by 43
Pseudomonas sp.

43
6 FT-IR absorption spectrum of EPS produced by
Microbacterium resistens
44
FT-IR absorption spectrum of EPS produced by
7
Bacillus cereus

8 GC-MS analysis of EPS produced by Pseudomonas sp. 44