Efficiency of Genetic Transformation via Pollen-Tube Pathway of Jatropha (Jatropha curcas L.) Based on Histochemical and Molecular Analysis

The genetic transformation via pollen-tube pathway is an alternative method to overcome the constraints imposed by genotype specificity in transformation and regeneration in jatropha (Jatropha curcas L.) tissue culture. Therefore, it is necessary to establish important parameters for efficient genetic transformation of jatropha via pollen-tube pathway. The objective of the research was to study the efficiency of direct transformation of jatropha via pollen-tube pathway based on histochemical and molecular analysis. Solution of purified pCAMBIA1301 DNA plasmid carrying a hptII marker gene and a gus reporter gene with concentration level of 0.05, 0.25, 0.50 µg µl-1 were applied to stigma of flowers at 1, 2, 4, 7, 10 h after pollination. Seedling of IP3A, IP3P and JcUMM18 jatropha’s genotypes derived from 15 combination treatments of plasmid DNA concentration and application time, also wild type was subjected to histochemical and molecular analyses. Based on those analyses, the efficiency of transformation via pollen-tube pathway of three jatropha genotypes ranged from 1.5-16.7%. PCR analysis showed that a number of positive plants were identified by using specific primers hptII and gus, i.e. 1-3 and 3-7 plants of the 15 combined treatments, respectively. It indicated that the transformation efficiency via the pollen-tube pathway varied in each jatropha genotype. Keywords: Jatropha curcas L., pCAMBIA1301, plasmid DNA, stigma-drip

[1]  M. Madhaiyan,et al.  Leaf-residing Methylobacterium species fix nitrogen and promote biomass and seed production in Jatropha curcas , 2015, Biotechnology for Biofuels.

[2]  K. Akashi,et al.  Efficient genetic transformation of Jatropha curcas L. by means of vacuum infiltration combined with filter-paper wicks , 2015, In Vitro Cellular & Developmental Biology - Plant.

[3]  H. Mao,et al.  Production of marker-free transgenic Jatropha curcas expressing hybrid Bacillus thuringiensis δ-endotoxin Cry1Ab/1Ac for resistance to larvae of tortrix moth (Archips micaceanus) , 2014, Biotechnology for Biofuels.

[4]  Xuede Wang,et al.  An efficient and highly reproducible approach for the selection of upland transgenic cotton produced by pollen tube pathway method , 2013 .

[5]  Baohong Zhang,et al.  Cotton transformation via pollen tube pathway. , 2013, Methods in molecular biology.

[6]  Hideyuki Suzuki,et al.  Development of transgenic plants in jatropha with drought tolerance , 2012 .

[7]  Hongzi Cong,et al.  Antisense suppression of alcohol acetyltransferase gene in ripening melon fruit alters volatile composition , 2012 .

[8]  W. Xiaojuan,et al.  Grain quality evaluations of hybrids between Triticum aestivum and Sorghum bicolor produced by pollen tube pathway method. , 2012 .

[9]  J. Aarrouf,et al.  Histological study of organogenesis in Cucumis melo L. after genetic transformation: why is it difficult to obtain transgenic plants? , 2011, Plant Cell Reports.

[10]  K. Fukui,et al.  Physical wounding-assisted Agrobacterium-mediated transformation of juvenile cotyledons of a biodiesel-producing plant, Jatropha curcas L. , 2011, Plant Biotechnology Reports.

[11]  B. Jha,et al.  Efficient genetic transformation of Jatropha curcas L. by microprojectile bombardment using embryo axes. , 2011 .

[12]  K. Fukui,et al.  Adventitious shoot regeneration from juvenile cotyledons of a biodiesel producing Plant, Jatropha curcas L. , 2011, Journal of bioscience and bioengineering.

[13]  T. Radhakrishnan,et al.  Stable genetic transformation of Jatropha curcas via Agrobacterium tumefaciens-mediated gene transfer using leaf explants , 2010 .

[14]  Liquan Zhang,et al.  Transformation of a marker-free and vector-free antisense ACC oxidase gene cassette into melon via the pollen-tube pathway , 2010, Biotechnology Letters.

[15]  R. Vicentini,et al.  Transcriptome analysis of the oil-rich seed of the bioenergy crop Jatropha curcas L , 2010, BMC Genomics.

[16]  K. Xiao,et al.  Establishment of transgenic cotton lines with high efficiency via pollen-tube pathway , 2009 .

[17]  Dharman Dhakshanamoorthy,et al.  EXTRACTION OF GENOMIC DNA FROM JATROPHA sp. USING MODIFIED CTAB METHOD , 2009 .

[18]  K. Rao,et al.  Tissue Culture-independent In Planta Transformation Strategy: an Agrobacterium tumefaciens-Mediated Gene Transfer Method to Overcome Recalcitrance in Cotton (Gossypium hirsutum L.) , 2008 .

[19]  Meiru Li,et al.  Establishment of an Agrobacteriuim-mediated cotyledon disc transformation method for Jatropha curcas , 2008, Plant Cell, Tissue and Organ Culture.

[20]  R. Jefferson.  Assaying chimeric genes in plants: The GUS gene fusion system , 1987, Plant Molecular Biology Reporter.

[21]  S. Riazuddin,et al.  Screening for putative transgenic rice and cotton plants: A simple and Easy Method , 2000 .