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RISS 인기검색어
- 검색키워드
- 저자 : Kathryn Kamo (검색결과 5 건)
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Biolistic-mediated Transformation of Lilium longiflorum cv. Nellie White
Kamo, Kathryn,Han, Bong Hee American Society for Horticultural Science 2008 HortScience Vol.43 No.6
<P>Slow-growing compact calluses were initiated from bulb scales of <I>Lilium longiflorum</I> cv. Nellie White that had been cultured for at least 6 months on Murashige and Skoog (MS) medium with 9 μm dicamba. To develop a reliable selection system, the sensitivity of nontransformed calluses and in vitro plants to different selective agents such as phosphinothricin, kanamycin, geneticin, paromomycin, and hygromycin was tested when grown on MS medium. Nontransformed calluses showed high sensitivity to 0.5 mg·L<SUP>−1</SUP> phosphinothricin, 25 mg·L<SUP>−1</SUP> geneticin, and 5 mg·L<SUP>−1</SUP> hygromycin. Nontransformed plants grown in vitro died on either 2 mg·L<SUP>−1</SUP> phosphinothricin or 75 mg·L<SUP>−1</SUP> hygromycin. Plants did not die when grown on either 200 mg·L<SUP>−1</SUP> kanamycin or 100 mg·L<SUP>−1</SUP> geneticin, and 100 mg·L<SUP>−1</SUP> paromomycin stimulated plant growth. Transformation was achieved using biolistics on callus bombarded with either the <I>bar-uidA</I> fusion gene under control of the CaMV 35S promoter or <I>npt II</I> and <I>uidA</I> under control of the CaMV 35S promoter. One week after biolistic bombardment, callus bombarded with the <I>bar-uidA</I> fusion gene was cultured for 1 month on MS medium supplemented with 9 μm dicamba and 0.1 mg·L<SUP>−1</SUP> phosphinothricin and then transferred to 0.2 mg·L<SUP>−1</SUP> phosphinothricin for 1 month followed by 1.0 mg·L<SUP>−1</SUP> for the next 4 months. Regenerating shoots and well-established plants were cultured on MS medium lacking hormones and with either 0.2 mg·L<SUP>−1</SUP> or 2.0 mg·L<SUP>−1</SUP> phosphinothricin, respectively. Callus bombarded with the <I>npt II</I> gene was cultured on MS medium with 50 mg·L<SUP>−1</SUP> geneticin until shoots regenerated. Regenerated shoots were cultured on MS medium lacking hormones. Under optimal conditions, 10 transgenic plants were selected from seven plates of callus bombarded with the <I>bar-uidA</I> fusion gene using phosphinothricin for selection. Both Southern hybridization of genomic DNA and polymerase chain reaction analysis verified the presence of the transgene in transformed ‘Nellie White’ plants. Transgenic plants were phenotypically normal, and they were crossed with nontransformed plants of <I>L. longiflorum</I> cvs. Sakai, Yin tung, Sakai, and Flavo. The presence of the <I>bar</I> gene in 41% of the T1 progeny plants confirmed stable integration of the transgene into the genomic DNA of transgenic lily plants. β-glucuronidase expression resulting from the <I>uidA</I> gene was demonstrated in leaves and roots of some of the transgenic lily plants by histochemical staining, determination of the specific activity of the β-glucuronidase enzyme, and Northern hybridization.</P>
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Vitrification of Gladiolus Shoot Tips from Cormels
Hyang Young Joung,Maria Cantor,David Ellis,Kathryn Kamo 한국원예학회 2007 Horticulture, Environment, and Biotechnology Vol.48 No.4
Gladiolus shoot tips, 1-2 ㎜, were excised from in vitro and greenhouse-grown cormels of cultivar ‘Peter Pears,’ in vitro-grown cormels of ‘Jenny Lee,’ field-grown cormels of the breeding lines 02-943A, 02-900, 02-926, and field-grown cormels of the cultivar ‘Double Deligh’. The major factors that affected growth of shoot tips following vitrification were size and dormancy of the excised shoot tip, cultivar, and time in Plant Vitrification Solution 2 (PVS2). The highest survivals of shoot tips was achieved using either greenhouse-grown ‘Peter Pears’ (77%) and field-grown 02-943A (55%). Recovery of viable shoot tips was best with incubation times of 30 and 60 minutes in PVS2 for ‘Jenny Lee’ and 02-943A, respectively. All shoots that have grown following vitrification were phenotypically normal in the greenhouse.
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Transgenic Plants of Easter Lily (Lilium longiflorum) with Phosphinothricin Resistance
Ahn, Byung Joon,Joung, Young Hee,Kamo, Kathryn K. The Korean Society of Plant Biotechnology 2004 Plant molecular biology and biotechnology research Vol.6 No.1
Transient uidA expression was used to optimize parameters required for biolistic transformation of suspension cells of Easter lily, Lilium longiflourm. Maximum uidA expression occurred following bombardment with gold particles as compared to tungsten. A 3hr pre-treatment of suspension cells with 0.125M osmoticum resulted in a 1.5X increase in uidA expression. A helium pressure of 1550 psi combined with a particle travelling distance of 6cm resulted in maximum uidA expression as compared to either 1100, 1200, or 1800 psi. Transient transformation resulted in up to 493 uidA expressing cells/Petri plate. For stable transformation suspension cells of Lilium longiflorum, were co-bombarded with plasmid DNA containing cucumber mosaic virus (CMV) replicase under the rice actin (Act1) promoter and either the bar or PAT genes under the cauliflower mosaic virus (CaMV 355) promoter. Ten regenerated plants contained the transgene as analyzed by PCR, and two of the ten plants were confirmed to contain the transgene by Southern hybridization. The two transgenic plants were independent transformants, one containing the bar gene and the other both the CMV replicase and bar genes. Plants were sprayed at the rosette stage and found to be resistant to 1000 mg/L of phosphinothricin (Trade name-Ignite) indicating expression of the bar gene throughout the leaves when bar was under control of the CaMV 35S promoter.
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