Molecular Biology Lab

Department of Biochemistry & Molecular Biology, University of Dhaka.
 
Current project: HEQEP AIF (WINDOW 2)
 
Creating a next generation of postgraduate researchers highly skilled in molecular and analytical approaches while harnessing endophytes for broad spectrum industrial application as well as using them for improved stress tolerance in jute.
 

Background of this project:

Plant associated microbes (generally bacteria and fungi) that live within plants as an integral part of the host metabolism and function are referred to as endophytes. Several recent studies have shown that this intrinsic microbial community brings significant benefits to plants. Endophytes have been shown to stimulate growth, increase nitrogen fixation, enhance protection against pathogens, increase drought resistance help the plant in obtaining nutrients and regulate phytohormones. However, this symbiotic mutualism can be species and environment specific. Identification and characterization of novel metabolites from endophytic microbiota may lead to the discovery of new drugs for human, plants and animals. Very few attempts have been undertaken to study their roles in the growth and metabolism of jute and allied fiber-producing plants. We aim to identify the fungal and bacterial endophytes of jute, determine their mutualistic relationship through biochemical, molecular biology, genomics and proteomics analyses, investigate potential industrial and medical applications through identifying important secondary metabolites and genes, assess their roles as mediators of biotic and abiotic stress resistance and growth enhancers as this important crop is being pushed towards inhospitable terrains in our country to make space for the food crops. This will open new windows for potential application of endophytes in improving growth and cultivation of jute, phytoremediation, organic farming, bio-control, accelerated retting, etc. We highly anticipate that these applications will be extended to other major crops.
 

Aims of the project:

Establishment of in vitro culture of jute endophytes, their characterization, and assessment of beneficial effects on jute growth and yield:
  • Application of beneficial endophytes in promoting growth, biotic and abiotic stress resistance, etc.
  • Application of endophytes for limiting plant pathogens will reduce the use of toxic chemicals, which will improve the quality of lives of the farmers and make the environment greener and reduce the negative impact on non-target species.
 
Characterization of some beneficial genes, proteins and secondary metabolites of jute endophytes:
  • Isolation of commercially and industrially important genes and proteins.
  • Applicability of medically and therapeutically important compounds isolated from endophytes.
 
Transcriptomic analysis of fungal endophyte of jute:
  • Assessment of broad spectrum interrelationship of endophytes with plants.
 
Use of endophytes for improving yield of jute varieties, and potentially other crops:
  • Identification of a combination of endophytes that provide maximum benefitsunder a number of different conditions.

 

What we have done

Development of jute EST/cDNA libraries and identification of genes of economic importance.
 

Objectives:


Identification of genes conferring stress tolerance in traditional jute varieties and use of these genes or markers linked to these traits to obtain stress tolerant jute suitable for growing in adverse conditions like drought, low temperature, salinity etc.
 

Our achievements:


1) Identification of gene(s) associated with low temperature tolerance in jute.

2) A complete gene sequence and mRNA sequence of a Putative Leucine Rich Transmembrane Protein Kinase has been deduced using degenerate PCR, RT-PCR, 3´ RACE and 5´ RACE. The expression of this putative Leucine Rich Transmembrane Protein Kinase has been also observed by Semi-quantitative PCR and Real Time PCR.

3) A total 2249 bp of a putative vps51/vps67 gene sequence has been deduced by using degenerate PCR, RT-PCR, 3´ RACE and 5´ RACE. The characterization of this putative gene is the present focus of our lab.

4) Non radioactive differential display identified a fragment of gene from low temperature tolerant jute (SDLT, Short Day Low Temperature) variety. The sequence of this putative gene has homology with sequences from prokaryotes only and bioinformatics studies suggest that this sequence contains a domain called EAL/GGDEF which has no homology with any of the eukaryotes sequenced till today. Expression of this gene has been studied by Semi-quantitative PCR and Real Time PCR and the presence of this gene in jute has been confirmed by Southern hybridization.

5) Development of efficient genetic transformation protocol of jute We successfully established a tissue culture independent transformation protocol for jute. In this approach young jute plants were transformed at shoot apical meristematic region using Agrobacterium tumefaciens. Heritable transmission of the transgene to progeny from genetically modified plants was confirmed by gus gene expression by histochemical analysis, selection on kanamycin containing medium, RT-PCR, PCR amplification and Southern hybridization. Efficiency of transformation was determined by selection on medium containing kanamycin, and inheritance of transgene to T2 generation plants.

6) DNA fingerprinting key for different jute genotypes was generated using jute specific SSR markers on 10 jute cultivars from two Corchorus species ( C. olitorius and C. capsularis) and the genetic relatedness among the cultivars was estimated.

7) Identification of SSR markers linked to mite tolerance in jute. SSR markers for jute have been successfully used in distinguishing mite sensitive and tolerance jute varieties. This marker has the potential of being useful in Marker Assisted Selection (MAS) in jute breeding programs for selection of lines resistant to mite.

8) Identification of genes conferring fungus resistance. Some differentially expressed genes have been amplified from fungus susceptible plants and not from the resistant ones. In other cases of differential expression, amplifications occurred only with samples at certain period of infection. Therefore, both constitutive and induced pattern of differential gene expression can be observed.