scholarly journals Transformation of Grape (Vitis vinifera L.)

HortScience ◽  
1995 ◽  
Vol 30 (4) ◽  
pp. 876F-876
Author(s):  
R. Scorza ◽  
J.M. Cordts ◽  
D.J. Gray ◽  
D.W. Ramming ◽  
R.L. Emershad

Transgenic grapevines were regenerated from somatic embryos produced from immature zygotic embryos of two seedless grape selections and from leaves of in vitro-grown plants of `Thompson Seedless'. Somatic embryos were bombarded with gold microparticles using the Biolistic PDS-1000/He device (Bio-Rad Labs) and then exposed to engineered A. tumefaciens EHA101 (E. Hood, WSU). Alternately, somatic embryos were exposed to A. tumefaciens without bombardment. Following cocultivation, secondary embryos multiplied on Emershad and Ramming proliferation medium under kan selection. Transgenic embryos were identified after 3 to 5 months and developed into rooted plants on woody plant medium with 1 mM N6-benzyladenine, 1.5% sucrose, and 0.3% activated charcoal. Seedless selections were transformed with plasmids pGA482GG (J. Slightom, Upjohn) and pCGN7314 (Calgene), which carry GUS and NPTII genes. `Thompson Seedless' was transformed with pGA482GG and pGA482GG/TomRSVcp-15 (D. Gonsalves, Cornell Univ.) containing the tomato ringspot virus coat protein gene. Integration of foreign genes into grapevines was verified by growth on kan, GUS, and PCR assays, and Southern analyses.

1996 ◽  
Vol 121 (4) ◽  
pp. 616-619 ◽  
Author(s):  
R. Scorza ◽  
J.M. Cordts ◽  
D.J. Gray ◽  
D. Gonsalves ◽  
R.L. Emershad ◽  
...  

Transgenic grape plants were regenerated from somatic embryos derived from leaves of in vitro-grown plants of `Thompson Seedless' grape (Vitis vinifera L.) plants. Somatic embryos were either exposed directly to engineered Agrobacterium tumefaciens or they were bombarded twice with 1-μm gold particles and then exposed to A. tumefaciens. Somatic embryos were transformed with either the lytic peptide Shiva-1 gene or the tomato ringspot virus (TomRSV) coat protein (CP) gene. After cocultivation, secondary embryos proliferated on Emershad/Ramming proliferation (ERP) medium for 6 weeks before selection on ERP medium containing 40 μg·mL-1 kanamycin (kan). Transgenic embryos were identified after 3 to 5 months under selection and allowed to germinate and develop into rooted plants on woody plant medium containing 1 μm 6-benzylaminopurine, 1.5% sucrose, 0.3% activated charcoal, and 0.75% agar. Integration of the foreign genes into these grapevines was verified by growth in the presence of kanamycin (kan), positive β-glucuronidase (GUS) and polymerase chain-reaction (PCR) assays, and Southern analysis.


HortScience ◽  
1995 ◽  
Vol 30 (4) ◽  
pp. 783E-783
Author(s):  
S.K. Dhir ◽  
U.L. Yadava

An efficient protocol has been developed for the in vitro multiplication of papaya (Carica papaya L.) through somatic embryogenesis utilizing immature zgotic embryos. Somatic embryos were initiated on MS basel media supplemented with 5 mg·liter–1 2,4-D, 400 mg·liter–1 glutamine, and 6% sucrose. After culturing for 2 months, 65% of the explants became highly embryogenic. Each explant produced 50 to 80 embryos in 4 months on culture induction medium. Frequency of embryogenesis was increased (75 to 150 somatic embryos on 80% explants) upon supplementing medium with 4% maltose as a carbon source and 100 mg·liter–1 L-asparagine. The embryogenic callus appeared yellow and embryos at different stages of development were well-organized. On regular subculturing, these cultures continued to produce secondary embryos. Following their transfer to the hormone-free medium supplemented with 4% maltose, these embryos germinated. The somatic embryogenesis system is rapid, repetitive, and highly proliferative. Thus, this system may have a potential use in the development of synthetic seed and transgenic papaya plants. Details of important factors affecting somatic embryogenesis will be discussed.


HortScience ◽  
1998 ◽  
Vol 33 (3) ◽  
pp. 557c-557
Author(s):  
Fred K. Westphal ◽  
Michael E. Compton

Torpedo-stage somatic embryos were selected from actively growing cultures and trasferred to embryo maintenance medium [MS with (per liter) 412.5 mg NH4NO3, 475 mg KNO3, 1 g myo-inositol, 90 g sucrose, 2 g activated charcoal, and 7 g TC agar] supplemented with either 0%, 2.5%, 5%, 7.5%, or 10% polyethylene glycol (PEG) for 4, 8, or 12 weeks. Embryos placed on treatment media were transferred directly to grape somatic embryo germination medium [MS with (per liter) 1 g myo-inositol, 30 g sucrose, 1 M benzyladenine, and 7 g TC agar] once their PEG treatment was terminated. The number of embryos that germinated was recorded 4 weeks after transfer to somatic embryo germination medium. The number of germinated embryos that differentiated into plants was recorded at 8 weeks. There was no difference in germination rates and embryo differentiation among embryos incubated on medium with or without PEG for 4 weeks. A difference in embryo growth rate was observed after 8 weeks on medium with PEG. Embryo grew fastest on media containing 5% or 7.5% PEG. In addition, embryos grown on medium with 5% or 7.5% PEG were morphologically similar to zygotic embryos.


2019 ◽  
Author(s):  
Jordan Demone ◽  
Jingqin Mao ◽  
Shen Wan ◽  
Maryam Nourimand ◽  
Äsbjörn Erik Hansen ◽  
...  

AbstractThe ‘triple-blue’ cultivar of blue spruce (Picea pungens Hoopsii) is notably recalcitrant towards the realm of traditional vegetative propagation methods. Its ability to naturally proliferate is limited by ovule and embryo abortion during the growing season, leading to low viable seed yield. In this study, we established a protocol using somatic embryogenesis (SE) as a means of propagating this popular ornamental cultivar. We collected cones from Hoopsii trees at seven different timepoints throughout the growing season (mid-June to late July in Ottawa (Plant Hardiness Zone 5A)). Female megagametophytes were harvested following each collection and immature zygotic embryos were plated onto induction media. Early somatic embryos began developing from the embryonic tissue (ET) three to five weeks following induction. The highest ET initiation frequency occurred from embryos collected June 20–July 10, suggesting that developmental stage of the embryo was a significant factor in SE induction. The conversion of mature somatic embryos into plantlets (emblings) was completed in eight–ten weeks at a rate of 92.8%. In this study, we demonstrate that in vitro somatic embryogenesis using our optimized protocol is a fast and prolific method for the mass propagation of Hoopsii blue spruce. This is the first report on the production of somatic Hoopsii emblings.


CORD ◽  
2017 ◽  
Vol 33 (2) ◽  
pp. 11
Author(s):  
Anitha Karun

Coconut is one of the principal crops of India cultivated in over 35 districts mainly in the southern states. The productivity of the crop is declining in many of the traditionally cultivated regions owing to ageing plantations as well as biotic and abiotic stresses. These plantations are to be replanted with high yielding varieties/hybrids for which adequate quantity of quality planting material is not available. Even though tissue culture research was initiated in many laboratories in the country, the work was eventually phased out in most of the laboratories for want of a repeatable protocol.  At ICAR-CPCRI, coconut tissue culture programs have been continuing for the past three decades. The attempts made include experimentation with different explants viz., immature inflorescence, plumular tissues, mature palm shoot meristem, ovary and anthers and different culture media supplemented with varying levels and types of hormones. Some of the successful protocols developed at the Institute include coconut zygotic embryo culture for collection and exchange of germplasm, cryopreservation and retrieval of zygotic embryos and pollen and plantlet regeneration from plumular tissues. Even though ICAR-CPCRI has succeeded in obtaining plantlets via direct organogenesis from inflorescence explants, the absence of friable calli formation from explants, the low rate of somatic embryo formation, large number of cultures turning to abnormal shoot development, non conversion of somatic embryos into plantlets, and formation of abnormal somatic embryos remain the major bottlenecks. Gene expression studies are being currently undertaken to decipher the molecular basis of in vitro recalcitrance.


2016 ◽  
Vol 74 (1) ◽  
Author(s):  
Pauline Destinugrainy KASI ◽  
. SUMARYONO

Summary In vitro culture of sago (Metroxylon sagu Rottb.) on an agar-solidified medium consists of somatic embryos of different sizes, colors, and developmental stages.  One gram of mostly globular somatic embryos were cultured on a solid medium to observe their morphological variations with respect to embryo size, color, and developmental stage over one passage of six weeks culture.  The medium was a modified-MS medium with half-strength of macronutrients containing   0.01 mg/L ABA and 2 mg/L kinetin.  At the end of culture passage, fresh weight of embryo increased by 2.3 folds.  The embryo numbers increased by more than two times indicating the formation of secondary embryos.  The average size of sago somatic embryos did not change significantly over the culture period; however, the embryo size was already highly varied at the start and increased gradually as the embryo developed.  At the initial of culture,   33.7 % of the embryos were yellowish, 64.1 % were greenish, and 2.2% were reddish.  By the end of the culture the composition of yellowish embryos increased to 51.2 %, greenish embryo decreased to 42.5 % and red embryos increased to 6.3 %.  At the initial culture, 61 % of the embryos were at the globular, 9 % at heart-shape and 30 % at torpedo stage.  Generally globular embryos developed into later-stage embryos as the culture progressed, although almost 56% of the embryos remained at the globular stage after the sixth week.Ringkasan Kultur in vitro sagu (Metroxylon sagu Rottb.) pada medium padat terdiri dari embrio somatik dalam berbagai ukuran, warna, dan fase perkembangan.  Satu gram embrio somatik yang sebagian besar dalam fase globuler dikulturkan pada medium padat untuk mengamati keragaman morfologi embrio dalam hal ukuran, warna dan fase perkembangan dalam satu periode kultur enam minggu.  Medium kultur adalah MS modifikasi dengan setengah hara makro serta penambahan zat pengatur tumbuh ABA 0,01 mg/L dan kinetin 2 mg/L.  Pada akhir masa kultur bobot embrio segar meningkat 2,3 kali dibandingkan awal masa kultur.  Jumlah embrio juga mengalami peningkatan sebesar lebih dari dua kali yang menunjukkan adanya pembentukan embrio somatik sekunder. Ukuran rata-rata embrio tidak berubah secara signifikan selama masa kultur akan tetapi ukuran embrio telah sangat beragam pada awal kultur dan terus meningkat hingga akhir kultur. Warna embrio mengalami perubahan selama periode kultur.  Pada awal kultur dijumpai 33,7 % embrio berwarna kuning, 64,1 % embrio hijau, dan 2,2 % embrio merah.  Pada akhir kultur presentase embrio kuning meningkat menjadi 51,2 %, embrio hijau menjadi 42,5 %, dan embrio merah 6,3 %.  Pada awal kultur, dijumpai 61 % embrio pada fase globuler, 9 % fase bentuk-hati dan 30 % fase torpedo.  Umumnya embrio globuler berkembang menjadi embrio fase lanjut selama kultur berlangsung, namun 56 % embrio masih tetap dalam fase globuler pada minggu keenam.


2016 ◽  
Vol 74 (1) ◽  
Author(s):  
Pauline Destinugrainy KASI ◽  
. SUMARYONO

Summary In vitro culture of sago (Metroxylon sagu Rottb.) on an agar-solidified medium consists of somatic embryos of different sizes, colors, and developmental stages.  One gram of mostly globular somatic embryos were cultured on a solid medium to observe their morphological variations with respect to embryo size, color, and developmental stage over one passage of six weeks culture.  The medium was a modified-MS medium with half-strength of macronutrients containing   0.01 mg/L ABA and 2 mg/L kinetin.  At the end of culture passage, fresh weight of embryo increased by 2.3 folds.  The embryo numbers increased by more than two times indicating the formation of secondary embryos.  The average size of sago somatic embryos did not change significantly over the culture period; however, the embryo size was already highly varied at the start and increased gradually as the embryo developed.  At the initial of culture,   33.7 % of the embryos were yellowish, 64.1 % were greenish, and 2.2% were reddish.  By the end of the culture the composition of yellowish embryos increased to 51.2 %, greenish embryo decreased to 42.5 % and red embryos increased to 6.3 %.  At the initial culture, 61 % of the embryos were at the globular, 9 % at heart-shape and 30 % at torpedo stage.  Generally globular embryos developed into later-stage embryos as the culture progressed, although almost 56% of the embryos remained at the globular stage after the sixth week.Ringkasan Kultur in vitro sagu (Metroxylon sagu Rottb.) pada medium padat terdiri dari embrio somatik dalam berbagai ukuran, warna, dan fase perkembangan.  Satu gram embrio somatik yang sebagian besar dalam fase globuler dikulturkan pada medium padat untuk mengamati keragaman morfologi embrio dalam hal ukuran, warna dan fase perkembangan dalam satu periode kultur enam minggu.  Medium kultur adalah MS modifikasi dengan setengah hara makro serta penambahan zat pengatur tumbuh ABA 0,01 mg/L dan kinetin 2 mg/L.  Pada akhir masa kultur bobot embrio segar meningkat 2,3 kali dibandingkan awal masa kultur.  Jumlah embrio juga mengalami peningkatan sebesar lebih dari dua kali yang menunjukkan adanya pembentukan embrio somatik sekunder. Ukuran rata-rata embrio tidak berubah secara signifikan selama masa kultur akan tetapi ukuran embrio telah sangat beragam pada awal kultur dan terus meningkat hingga akhir kultur. Warna embrio mengalami perubahan selama periode kultur.  Pada awal kultur dijumpai 33,7 % embrio berwarna kuning, 64,1 % embrio hijau, dan 2,2 % embrio merah.  Pada akhir kultur presentase embrio kuning meningkat menjadi 51,2 %, embrio hijau menjadi 42,5 %, dan embrio merah 6,3 %.  Pada awal kultur, dijumpai 61 % embrio pada fase globuler, 9 % fase bentuk-hati dan 30 % fase torpedo.  Umumnya embrio globuler berkembang menjadi embrio fase lanjut selama kultur berlangsung, namun 56 % embrio masih tetap dalam fase globuler pada minggu keenam.


Euphytica ◽  
2006 ◽  
Vol 152 (2) ◽  
pp. 217-224 ◽  
Author(s):  
X. M. Yang ◽  
Z. Y. Cao ◽  
L. Z. An ◽  
Y. M. Wang ◽  
X. W. Fang

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