High efficiency transformation protocol for three Indian cotton varieties via Agrobacterium tumefaciens

A protocol for consistent production of transgenic cotton plants in three Indian varieties was established utilizing Agrobacterium-mediated transformation. Shoot tip explants were transformed by cocultivation with Agrobacterium tumefaciens strain LBA 4404. The strain harbors a binary vector pBAL2 carrying the reporter gene $_\beta$-glucuronidase intron (GUS-INT) and the marker gene neomycin phosphotransferase (NPTII). Regeneration potential of explants or different hormones was studied in detail. Among the different combinations of BAP and NAA tested, 0.1 mg $1^{-1}$ of BAP and NAA in the medium influenced efficient regeneration of shoots by organogenesis. Shoot bud proliferation and elongation was achieved in 3�4 weeks time on medium supplemented with $GA_3$. The putatively transformed shoots were harvested and placed for rooting on medium containing IBA and $75mg \hspace{2mm} 1^{-1}$ kanamycin. Transgenic plants were recovered in 12�16 weeks from the time of gene transfer to establishment in pots. Molecular analysis of the field established plantlets was carried to confirm the transgenic nature. The presence of GUS and NPTII genes in the transgenic plants was verified by histochemical GUS assay and polymerase chain reaction (PCR) analysis, respectively. Integration of T-DNA into the genome of putative transgenics was further confirmed by Southern blot analysis. A total of 70�75 transgenic plants were raised in pots. Progeny analysis of these plants showed a classical Mendelian pattern of inheritance

Expression of hemagglutinin protein of Rinderpest virus in transgenic pigeon pea [Cajanus cajan (L.) Millsp.] plants

Rinderpest virus is the causative agent of a devastating, often fatal disease in wild and domestic bovids that is endemic in Africa, the Middle East and South Asia. The existing live attenuated vaccine is heat-labile, and thus there is a need for the development of new strategies for vaccination. This paper reports the development of transgenic pigeon pea [Cajanus cajun (L.) Millsp.] expressing one of the protective antigens, the hemagglutinin (H) protein of Rinderpest virus. A 2-kb fragment containing the coding region of the H protein was cloned into pBI121 and mobilized into Agrobacterium tumefaciens strain EHA105. Embryonic axes and cotyledonary nodes from germinated seeds of pigeon pea were used for transformation. The presence of the transgene in transgenic plants was confirmed by Southern blots, and the specific transcription of the marker gene in the plants was demonstrated by reverse transcription-polymerase chain reaction. Integration of the H gene into the pigeon pea genome was confirmed by Southern hybridization. The expression of the H protein in the transgenic lines was confirmed by Western blot analysis using a polyclonal monospecific antibody to the H protein. The highest level of expression of the hemagglutinin protein in leaves of pigeon pea was 0.49% of the total soluble protein. The transgenic plants were fertile and the transgene expressed in the progeny

Development of cotton transgenics with antisense AV2 gene for resistance against cotton leaf curl virus (CLCuD) via Agrobacterium tumefaciens

Cotton transgenics for resistance against cotton leaf curl disease using antisense movement protein gene (AV2) were developed in an Indian variety (F846) via Agrobacterium-mediated transformation using the protocol developed previously. A binary vector pPZP carrying the antisense AV2 (350 bp) gene along with the nptII gene was used. Transgenic nature of the putative transgenics was confirmed by molecular analysis. Shoots were induced on selection medium and subcultured on rooting medium contg. IBA and $75 mg \hspace {3mm} l^ {-1}$ kanamycin. Transgenic plants were recovered in 12-16 weeks from the time of gene transfer to establishment in pots. Preliminary analysis of the field-established plantlets was conducted by PCR. $T_1$ plants were obtained from $T_0$ seeds, the presence of the AV2 and nptII genes in the transgenic plants was verified by PCR and integration of T-DNA with AV2 into the plant genome of putative transgenics was further confirmed by Southern blot analysis. Several $T_1$ lines were maintained in the greenhouse. Progeny anal. of these plants by PCR analysis showed a classical Mendelian pattern of inheritance

Expression of biologically active Hemagglutinin-neuraminidase protein of Peste des petits ruminants virus in transgenic pigeonpea [Cajanus cajan (L) Millsp.]

Hemagglutinin-neuraminidase (HN) gene of Peste des petits ruminants virus (PPRV) has been expressed in pigeonpea for the development of an edible vaccine for Peste des Petits ruminant (PPR). PPRV, causes PPR disease in sheep and goats with high mortality rate. The two surface glycoproteins of PPRV Hemagglutinin-neuraminidase and fusion protein (F) confer protective immunity. We report the successful generation of transgenic pigeonpea (Cajanus cajan (L.) Millsp.) plants and expression of HN protein having biological activity. A 2 fragment containing the coding region of the HN gene from an Indian isolate was cloned into the binary vector pBI121 and mobilized into Agrobacterium tumefaciens strain GV3 101. Cotyledonary nodes from germinated seeds of pigeonpea were used for transformation. The presence of transgenes, NPTII and HN in the plants was confirmed by PCR. The expression of HN protein in the transgenic lines was further confirmed by Western blot analysis using polyclonal monospecific antibody to HN and more importantly plant-derived HN protein was shown to be biologically active as demonstrated by neuraminidase activity. Transgenic plants were fertile and PCR of $T_1$ plants confirmed the inheritance of the transgene

Additional file 3: Table S2. of Discovery and mapping of genomic regions governing economically important traits of Basmati rice

Chi square values of microsatellite markers showing segregation distortion among F2 population of Basmati370/Jaya (DOC 154 kb

Additional file 10: Table S7 of Discovery and mapping of genomic regions governing economically important traits of Basmati rice

The genes with non-synonymous SNPs in the QTL for chalkiness qCHK4.1. (RM564-RM348) (DOC 168 kb

Additional file 9: Table S6 of Discovery and mapping of genomic regions governing economically important traits of Basmati rice

The genes with non-synonymous SNPs in the QTL for filled grain qFG1.1. (RM11968-RM14). (DOC 65 kb