productivity of guar and guar gum G. S. Randhawa Department of Biotechnology IIT Roorkee
Jan 03, 2016
Increasing productivity of guar and guar
gum
G. S. RandhawaDepartment of Biotechnology
IIT Roorkee
G. S. RandhawaDepartment of Biotechnology
IIT Roorkee
GUAR (CLUSTER BEAN)
A NEGLECTED MIRACLE BEAN
THE ECONOMIC TIMES
“Guar gum has overtook traditional heavyweights of India’s top farm export; rice and cotton, and seems to make its place into the league of top ten shipments of the country”
http://articles.economictimes.indiatimes.com/keyword/guar-gum (March 2013)
A strong need to enhance the productivity of guar crop and guar gum
BUT HOW ???
APPROACHES TO ENHANCE THE PRODUCTIVITY
Increasing area under cultivation
Improving agronomic practices
Controlling pests and diseases
By developing varieties having the following characterstics:
High yield
Stress tolerance
Disease and pest resistance
Improved gum quantity and quality
PROBLEMS IN ENHANCING PRODUCTIVITY
• Poor research funding
• Less number of scientists
• Poor infrastructure facilities
RESEARCH APPROACHES FOR IMPROVEMENT
• Classical / traditional approaches
• Biotechnological approaches
CLASSICAL/ TRADITIONAL APPROACHES • Germplasm collection
• Screening of germplasm
• Hybridization and selection
BIOTECHNOLOGICAL APPROACHES
• Molecular markers
• Identification of genes
• Characterization of guar proteins
• Regulation of gene expression
• Genetic manipulation
• Cell and tissue culture
• Sequencing of guar genome
MOLECULAR MARKERS
A fragment of DNA that is associated with a certain location within the genome is called molecular marker
Various types of molecular markers: RFLP, RAPD, AFLP, ISSR, SSR, SNP
Studies in guar:
RAPD (Liu et al., 2009, Punia et al., 2009 and Nagesh et al., 2013)
ISSR (Nagesh et al., 2013)
SSR (Nagesh et al., 2014)
Nagesh et al., 2013
Molecular characterization of cluster bean (Cyamopsis tetragonoloba L. Taub) using RAPD and ISSR markers
A total of 907 SSR-containing sequences were identified
Out of 224 primer pairs, 3 resulted in the amplification of SSR-containing sequences producing reliable and reproducible DNA bands in Cyamopsis tetragonoloba, C. serrata, and C. senegalensis, respectively
Identification and Characterization of EST-SSR Markers
Nagesh et al., 2014
Banding pattrens obtained from amplification of SSR markers
UDP-glucose
Pathway to galactomannan formation in plantsfructose-6-phosphate
phosphomannoisomerase (PMI)
mannose-6-phosphate
mannose-1-phosphate
mutase
PPi
GTPPPase
2Pi
GDP-mannose
GDP-mannose pyrophosphorylase
UDP-galactose
epimerase
-mannan synthase
-man-man-man-man--man-man-man-man-
gal
gal-galactosyltransferase
UDP
sucrose
UDP
fructose
hexok
inas
e
-mannan
galactomannan KSD/
020503
IDENTIFICATION OF GENES
Mannan synthase (Dhugga et al., 2004)
Galactomannan galactosyltransferase (Edwards et al., 1999)
α-galactosidase (Reid and Meier, 1973)
Genes involved in gum synthesis
Genes responsible for stress tolerance
?????
Genes responsible for plant growth and development
?????
ß-MANNAN SYNTHASE
The first plant cell wall hemicellulosic gene cloned
Dhugga et al., Science, 2004
Dr. Kanwarpal Singh Dhugga who cloned ManS gene of guar
Pioneer Hi-Bred International Inc. where ManS gene of guar was cloned
Peptide mass fingerprinting of a guar protein (Unpublished data from our lab)
In our lab cloning, expression, purification and characterizationof guar plant proteins is in progress
Studying of the pathway responsible for stress tolerance in guar using the physiological, biochemical and molecular approaches is being carried out in our lab. Identification of genes responsible for stress tolerance is in progress.
(Unpublished data)
Genes responsible for stress tolerance in guar
POTENTIAL STRENGTH OF PROMOTERSAnalysis of GUS expression in different tissues of transgenic alfalfa plants carrying GUS reporter genes under control of the MS, Phas and 35S promoters. Promoter activity was evaluated in roots, leaf, flowers, pods, seed coat, endosperm and embryos, respectively: a, d, g, j, m, p, s 35S promoter b, e, h, k, n, q, t MS promoter c, f, i, l, o, r, u b-phaseolin promoter
Naoumkina and Dixon, 2011
Characterization of mannan synthase promoter from guar
REGULATION OF GENE EXPRESSION
GENETICALLY MODIFIED PLANTS
In vivo modification of cell wall polysaccharide galactomannan of guar transformed with alpha-galactosidase gene cloned from senna
Sequence of Senna alpha galactosidase and Restriction enzyme map of the plant transformation vector, pBS-GNG-GAL, used to transform guar.
Joersbo et al. 2001
Transformed guar plant showing expression of reporter gus gene in leaf tissue(Unpublished data)
In our lab work on RNAi in guar is in progress
Genetically Modified Guar Plantlets(Unpublished data)
Explants used:1.Cotyledon
2.Hypocotyl
3.Cotyledonary node
(from in vitro germinated seedlings)
Different combinations of plant growth hormones used:For regeneration:Different concentrations of NAA -BAP, NAA-KN, IAA-KN, IAA-BAP, 2,4-D-BAP and 2,4-D-KN combinations
For elongation of regenerated shoots:Different concentrations of BA-IAA-GA3 in combination
For rooting:IAA and IBA alone
PLANT CELL AND TISSUE CULTURE
Verma et al., 2013
In vitro regeneration of complete plantlets in a single culture in guar
Verma et al., 2013
OUR TEAM
Nagesh K A
Manisha Choudhary
Swati VermaPranita Bhatele
Umesh K. Tanwar Shalini Pareek
Kulvinder S. Gill Kanwarpal S. Dhugga
Pallavi Gahlot
Navneet Kaur Deepa Dewan Nishu Mittal Poonam Jaiswal
Shilpi Kumari
Prof. S.K. Tripathi
Prof. S.M. Sondhi
Prof. Pravindra Kumar
Sardar Tara Singh JiSardar Tara Singh JiMy MentorMy Mentor
THANK YOU
My website WWW.GSRANDHAWA.IN