scholarly journals (283) Biolistic Genetic Transformation of Carica papaya L. using the Helios™ Gene Gun

HortScience ◽  
2006 ◽  
Vol 41 (4) ◽  
pp. 1053A-1053
Author(s):  
Salvador Guzmán-González ◽  
Pedro Valadez-Ramírez ◽  
Rosa-Edith Robles-Berber ◽  
Laura Silva-Rosales ◽  
José-Luis Cabrera-Ponce
Keyword(s):  
Genetic Transformation ◽  
Particle Bombardment ◽  
Carica Papaya ◽  
Resistance Mechanism ◽  
35S Promoter ◽  
Helium Pressure ◽  
Gene Gun ◽  
Gold Particle Size ◽  
Key Factor ◽  
Gus Transient Expression

Biolistic genetic transformation of plants with viral genes is a method for controlling plant virus diseases; however, optimization of the particle bombardment parameters according to the transformation system is a key factor for an appropiate transgene expression and, therefore, a stronger resistance mechanism in transgenic plants. In order to optimize biolistic parameters, somatic papaya (Carica papaya L.) cv. Maradol embryo masses were bombarded with the CAMBIA 1301 plasmid construction that contains the coat protein gene (CP) of the papaya ringspot virus isolate of Colima, Mexico, driven by the double constitutively CaMV 35S promoter and flanked for the GUS and hygromycin (hpt) resistance genes. Particle bombardment protocol was carried out using the Helios™ Gene Gun device (BioRad) and the manufacturer's instruction manual. Helium pressure (50, 100, and 150 psi) and gold particle size (0.6, 1.0, and 1.6 μm) were evaluated. Five days after bombardment, somatic embryo clusters were used for GUS transient expression and, during 2 months, were selected into 50, 75, and 150 mg·L-1 hygromycin-containing media to its later CP-PCR detection. Results showed that 50 psi and 1.0 μm were the two optimal values for the assayed analyses. This is the first report of genetic transformation of papaya using the Helios™ Gene Gun device as a new tool compared to conventional PDS-1000/He.

scholarly journals Genetic Transformation ofMetroxylon sagu(Rottb.) Cultures viaAgrobacterium-Mediated and Particle Bombardment

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Evra Raunie Ibrahim ◽  
Md. Anowar Hossain ◽  
Hairul Azman Roslan
Keyword(s):  
Genetic Transformation ◽  
Starch Content ◽  
Pcr Amplification ◽  
Culture Method ◽  
Helium Pressure ◽  
Gene Gun ◽  
Dot Blot ◽  
Suitable Material ◽  
Perennial Plant ◽  
Sago Palm

Sago palm (Metroxylon sagu) is a perennial plant native to Southeast Asia and exploited mainly for the starch content in its trunk. Genetic improvement of sago palm is extremely slow when compared to other annual starch crops. Urgent attention is needed to improve the sago palm planting material and can be achieved through nonconventional methods. We have previously developed a tissue culture method for sago palm, which is used to provide the planting materials and to develop a genetic transformation procedure. Here, we report the genetic transformation of sago embryonic callus derived from suspension culture usingAgrobacterium tumefaciensand gene gun systems. The transformed embryoids cells were selected against Basta (concentration 10 to 30 mg/L). Evidence of foreign genes integration and function of thebarandgusgenes were verified via gene specific PCR amplification, gus staining, and dot blot analysis. This study showed that the embryogenic callus was the most suitable material for transformation as compared to the fine callus, embryoid stage, and initiated shoots. The gene gun transformation showed higher transformation efficiency than the ones transformed usingAgrobacteriumwhen targets were bombarded once or twice using 280 psi of helium pressure at 6 to 8 cm distance.

Optimization of particle bombardment parameters for DNA delivery into the male flowers of banana

Biologia ◽  
2014 ◽  
Vol 69 (7) ◽  
Author(s):  
Fatemeh Mahdavi ◽  
Maziah Mahmood ◽  
Normah Noor
Keyword(s):  
Particle Bombardment ◽  
Fluorescent Protein ◽  
Constitutive Expression ◽  
Vacuum Pressure ◽  
35S Promoter ◽  
Helium Pressure ◽  
Gfp Expression ◽  
Gfp Gene ◽  
Male Flowers ◽  
Green Fluorescent

AbstractThe possibility of increasing the efficiency of banana transformation was investigated by particle bombardment of the male flowers of banana plants for constitutive expression of gfp gene. The effects of particle bombardment parameters, such as acceleration pressure, bombardment distance, chamber vacuum pressure, gold microcarrier size, gold quantity, DNA quantity, number of bombardments and pre-culture were examined. Single cauliflower-like bodies (CLBs) clusters, induced from meristemic parts of Musa sapientum cv. Nangka (AAB) male flowers, were bombarded by pCambia1304 plasmid carrying gfp gene driven by the CaMV 35S promoter. Optimal transient expression of green-fluorescent protein (GFP) was obtained when the three-day old cultured tissues were bombarded two times at 1100 psi helium pressure. However, the highest GFP expression was observed when 9 cm was applied as bombardment distance with 28 mmHg chamber vacuum pressure. Gold particle with 1 μm diameter at 60 μg/μL concentrations coated with 1.5 μg/μL of DNA have been used as the optimum bombardment parameter since GFP expression was significantly different compared to other conditions. Application of optimized condition proved effective for the generation of stable transgenic banana plants. PCR and southern blot analyses confirmed the presence and integration of gfp gene in genomic DNA of transformed plants. Transformation frequency achieved with the optimized protocol was 7.5% which was significantly higher than the conventional protocol.

scholarly journals Factors Affecting Transient Gene Expression in Mango Tissue Using Particle Bombardment

HortScience ◽  
2004 ◽  
Vol 39 (4) ◽  
pp. 754E-755
Author(s):  
Vaughan Agrez* ◽  
Zora Singh ◽  
Mis Gill
Keyword(s):  
Gene Expression ◽  
Particle Bombardment ◽  
Mangifera Indica ◽  
Transient Gene Expression ◽  
Gus Expression ◽  
35S Promoter ◽  
Helium Pressure ◽  
Factors Affecting ◽  
Pre Treatment

Particle bombardment seems to be a promising method for genetic transformation of mango and the information on various factors affecting transient gene expression is inconclusive. Pro-embryonic masses (PEMs) of mango (Mangifera indica L. cv. Kensington Pride) used for particle bombardment were established and maintained in vitro using various liquid and semisolid media containing 2,4-D. Various factors affecting the biolistic transformation efficiency in mango and subsequent transient gene expression were optimized using β-glucuronidase (GUS) as a reporter gene driven by CaMV 35S promoter. Our results show that bombardment pressure significantly affects transient gene expression with the best results obtained from PEMs bombarded at 1200 kPa of helium pressure under vacuum. The application of osmoticum pre and post-bombardment also significantly increased the transient gene expression in the PEMs as compared to the controls. Mannitol (0.2 m) proved to be the best osmoticum in improving transient GUS expression as compared to sorbitol. The duration of exposure of PEMs to mannitol (0.2 m) both pre and post-bombardment also played a significant role in improving transient gene expression. The transient GUS expression was significantly highest with a pre-treatment of 0.2 m mannitol for 4 hours as compared to 0, 8 and 12 hours. The post-bombardment treatment of 0.2 m mannitol for 16 hours resulted in significantly highest transient gene expression as compared to 0, 4, 8, 12, 20 and 24 hours. In conclusion, PEMs of `Kensington Pride' mango bombarded at 1200 kPa, which were exposed to mannitol (0.2 m) for 4 and 16 hours pre and post bombardment respectively, resulted in the highest transient GUS expression (25.1 GUS foci/mg PEMs).

scholarly journals Expression Analysis of Hairpin RNA CarryingSugarcane mosaic virus(SCMV) Derived Sequences and Transgenic Resistance Development in a Model Rice Plant

2017 ◽  
Vol 2017 ◽  
pp. 1-10 ◽  
Author(s):  
Sehrish Akbar ◽  
Muhammad Tahir ◽  
Ming-Bo Wang ◽  
Qing Liu
Keyword(s):  
Mosaic Virus ◽  
Particle Bombardment ◽  
Rice Plant ◽  
35S Promoter ◽  
Small Interfering Rnas ◽  
Transgenic Resistance ◽  
Hairpin Rna ◽  
Rice Callus ◽  
Transgenic Expression ◽  
Callus Tissues

Developing transgenic resistance in monocotyledonous crops against pathogens remains a challenging area of research.Sugarcane mosaic virus(SCMV) is a serious pathogen of many monocotyledonous crops including sugarcane. The objective of present study was to analyze transgenic expression of hairpin RNA (hpRNA), targeting simultaneouslyCP(Coat Protein) andHc-Pro(helper component-proteinase) genes of SCMV, in a model rice plant. Conserved nucleotide sequences, exclusive for DAG (Aspartic acid-Alanine-Glycine) and KITC (Lycine-Isoleucine-Threonine-Cysteine) motifs, derived from SCMVCPandHc-Progenes, respectively, were fused together and assembled into the hpRNA cassette under maize ubiquitin promoter to form Ubi-hpCP:Hc-Pro construct. The same CP:Hc-Pro sequence was fused with theβ-glucuronidase gene (GUS) at the 3′ end under CaMV 35S promoter to develop 35S-GUS:CP:Hc-Pro served as a target reporter gene construct. When delivered into rice callus tissues by particle bombardment, the Ubi-hpCP:Hc-Pro construct induced strong silencing of 35S-GUS:CP:Hc-Pro. Transgenic rice plants, containing Ubi-hpCP:Hc-Pro construct, expressed high level of 21–24 nt small interfering RNAs, which induced specific suppression against GUS:CP:Hc-Pro delivered by particle bombardment and conferred strong resistance to mechanically inoculated SCMV. It is concluded that fusion hpRNA approach is an affordable method for developing resistance against SCMV in model rice plant and it could confer SCMV resistance when transformed into sugarcane.

An efficient particle bombardment system for the genetic transformation of asparagus (Asparagus officinalis L.)

1997 ◽  
Vol 16 (5) ◽  
pp. 255-260 ◽  
Author(s):  
Jose Luis Cabrera-Ponce ◽  
Liliana L�pez ◽  
Nacyra Assad-Garcia ◽  
Consuelo Medina-Arevalo ◽  
Ana Maria Bailey ◽  
...  
Keyword(s):  
Genetic Transformation ◽  
Particle Bombardment ◽  
Asparagus Officinalis
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