Extraction of DNA from plant tissues

Rogers, Scott O.; Bendich, Arnold J.

doi:10.1007/978-94-017-5294-7_6

Cited by 497 publications

(423 citation statements)

References 7 publications

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“…The total plant genomic DNA was extracted from fresh young leaves of putative transformed (T 0 ) and non-transformed (control) plants by the CTAB (cetyl trimethyl ammonium bromide) method (Rogers and Bendich 1988).…”

Section: Dna Extractionmentioning

confidence: 99%

Development of an efficient in vitro plant regeneration system amenable to Agrobacterium- mediated transformation of a recalcitrant grain legume blackgram (Vigna mungo L. Hepper)

Sainger

Chaudhary

Dahiya

et al. 2015

Physiol Mol Biol Plants

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An efficient, rapid and direct multiple shoot regeneration system amenable to Agrobacterium-mediated transformation from primary leaf with intact petiole of blackgram (Vigna mungo) is established for the first time. The effect of the explant type and its age, type and concentration of cytokinin and auxin either alone or in combination and genotype on multiple shoot regeneration efficiency and frequency was optimized. The primary leaf explants with petiole excised from 4-day-old seedlings directly developed multiple shoots (an average of 10 shoots/ explant) from the cut ends of the petiole in 95 % of the cultures on MSB (MS salts and B 5 vitamins) medium containing 1.0 μM 6-benzylaminopurine. Elongated (2-3 cm) shoots were rooted on MSB medium with 2.5 μM indole-butyric acid and resulted plantlets were hardened and established in soil, where they resumed growth and reached maturity with normal seed set. The regenerated plants were morphologically similar to seed-raised plants and required 8 weeks time from initiation of culture to establish them in soil. The regeneration competent cells present at the cut ends of petiole are fully exposed and are, thus, easily accessible to Agrobacterium, making this plant regeneration protocol amenable for the production of transgenic plants. The protocol was further successfully used to develop fertile transgenic plants of blackgram using Agrobacterium tumefaciens strain EHA 105 carrying a binary vector pCAMBIA2301 that contains a neomycin phosphotransferase gene (nptII) and a β-glucuronidase (GUS) gene (uidA) interrupted with an intron.The presence and integration of transgenes in putative T 0 plants were confirmed by polymerase chain reaction (PCR) and Southern blot hybridization, respectively. The transgenes were inherited in Mendelian fashion in T 1 progeny and a transformation frequency of 1.3 % was obtained. This protocol can be effectively used for transferring new traits in blackgram and other legumes for their quantitative and qualitative improvements.

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Section: Dna Extractionmentioning

confidence: 99%

Development of an efficient in vitro plant regeneration system amenable to Agrobacterium- mediated transformation of a recalcitrant grain legume blackgram (Vigna mungo L. Hepper)

Sainger

Chaudhary

Dahiya

et al. 2015

Physiol Mol Biol Plants

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“…Total genomic DNA was extracted from nontransformed (control) and putative transformed Lemna fronds using a modified CTAB (cetyl trimethyl ammonium bromide) method (Roger and Bendich 1988) and used as template to amplify nptII and uidA genes by using primers specific to their coding regions. The polymerase chain reaction (PCR) was carried out with Taq DNA polymerase (Sigma, USA) in 25 μl reaction volume.…”

Section: Pcr Analysismentioning

confidence: 99%

Genetic transformation of Indian isolate of Lemna minor mediated by Agrobacterium tumefaciens and recovery of transgenic plants

Chhabra

Chaudhary

Sainger

et al. 2011

Physiol Mol Biol Plants

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Transgenic plants of an Indian isolate of Lemna minor have been developed for the first time using Agrobacterium tumefaciens and hard nodular cell masses 'nodular calli' developed on the BAP -pretreated daughter frond explants in B 5 medium containing sucrose (1.0 %) with 2,4-D (5.0 μM) and 2-iP (50.0 μM) or 2,4-D (50.0 μM) and TDZ (5.0 μM) under light conditions. These calli were co-cultured with A. tumefaciens strain EHA105 harboring a binary vector that contained genes for β-glucuronidase with intron and neomycin phosphortransferase. Transformed cells selected on kanamycin selection medium were regenerated into fronds whose transgenic nature was confirmed by histochemical assay for GUS activity, PCR analysis and Southern hybridization. The frequency of transformation obtained was 3.8 % and a period of 11-13 weeks was required from initiation of cultures from explants to fully grown transgenic fronds. The pretreatment of daughter fronds with BAP, use of non-ionic surfactant, presence of acetosyringone in co-cultivation medium, co-culture duration of 3 d and 16 h photoperiod during culture were found crucial for callus induction, frond regeneration and transformation of L. minor. This transformation system can be used for the production of pharmaceutically important protein and in bioremediation.

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“…Probe and block preparation: Total genomic DNA was extracted from the Longiflorum cultivar 'White Fox' and the Asiatic cultivar 'Polyanne' with the CTAB method (Rogers and Bendich 1988). 'White Fox' DNA was sonicated to 1-10 kb fragments and used as probe.…”

Section: Genomic In Situ Hybridizationmentioning

confidence: 99%

Genome composition of triploid lily cultivars derived from sexual polyploidization of Longiflorum × Asiatic hybrids (Lilium)

Zhou

Ramanna²,

Visser³

et al. 2007

Euphytica

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About 19 cultivars, which had originated from backcrosses between F1 LA (Longiflorum · Asiatic) hybrids (2n = 2x = 24) as female parents and Asiatic cultivars as male parents (2n = 2x = 24), were analyzed with genomic in situ hybridization. 17 of them were triploid (2n = 3x = 36), and two aneuploid (2n = 3x + 1 = 37). The triploid cultivars had resulted from the functional 2n eggs produced by the female parents (F1 hybrids) because first division restitution (FDR) occurred in their meiosis during megasporogenesis. Similarly, the aneuploid cultivars had originated from viable 2n + 1 eggs. The extra chromosome in cultivar 041555 or 041572 resulted from one univalent or one half-bivalent which might have lagged behind when the sister chromatids of the other univalents and half-bivalents were segregating during the FDR process in their LA hybrid parents, respectively. That the majority of cultivars possessed recombinant chromosomes showed that intergenomic recombination might play an important role during the selection of the cultivars directly from BC1 progenies. That five cultivars of the 15 recombinant cultivars only had reciprocal recombinant chromosomes and 10 cultivars had non-reciprocal recombinant chromosomes indicates that the latter are more important. Because 9 of the 10 non-reciprocal recombinant cultivars possessed substitutions for recombinant segments, it also indicated that such substitutions could be an important source for the genetic variation in the sexual triploid BC1 progenies. In such cases there was a potential for the expression of the recessive genes of the backcross parent in a nulliplex (aaa) condition in the substituted segments. Genetic variation resulting from such nulliplex loci might have played a role in the selection of some of the cultivars.

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Extraction of DNA from plant tissues

Cited by 497 publications

References 7 publications

Development of an efficient in vitro plant regeneration system amenable to Agrobacterium- mediated transformation of a recalcitrant grain legume blackgram (Vigna mungo L. Hepper)

Development of an efficient in vitro plant regeneration system amenable to Agrobacterium- mediated transformation of a recalcitrant grain legume blackgram (Vigna mungo L. Hepper)

Genetic transformation of Indian isolate of Lemna minor mediated by Agrobacterium tumefaciens and recovery of transgenic plants

Genome composition of triploid lily cultivars derived from sexual polyploidization of Longiflorum × Asiatic hybrids (Lilium)

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