Genetic transformation of Citrus sinensis cv. Hamlin with hrpN gene from Erwinia amylovora and evaluation of the transgenic lines for resistance to citrus canker

2009 ◽  
Vol 122 (1) ◽  
pp. 109-115 ◽  
Author(s):  
Janaynna Magalhães Barbosa-Mendes ◽  
Francisco de Assis Alves Mourão Filho ◽  
Armando Bergamin Filho ◽  
Ricardo Harakava ◽  
Steven V. Beer ◽  
...  
Keyword(s):  
Genetic Transformation ◽  
Citrus Sinensis ◽  
Erwinia Amylovora ◽  
Citrus Canker ◽  
Transgenic Lines

scholarly journals Attacin A Gene from Tricloplusia ni Reduces Susceptibility to Xanthomonas axonopodis pv. citri in Transgenic Citrus sinensis `Hamlin'

2006 ◽  
Vol 131 (4) ◽  
pp. 530-536 ◽  
Author(s):  
Raquel L. Boscariol ◽  
Mariza Monteiro ◽  
Elizabete K. Takahashi ◽  
Sabrina M. Chabregas ◽  
Maria Lucia C. Vieira ◽  
...  
Keyword(s):  
Citrus Sinensis ◽  
Fluorescent Protein ◽  
Blot Hybridization ◽  
Plasmid Vector ◽  
Southern Blot Hybridization ◽  
Citrus Canker ◽  
Green Fluorescent Protein Gene ◽  
Xanthomonas Axonopodis ◽  
Transgenic Lines

Citrus canker, caused by Xanthomonas axonopodis Starr and Garces pv. citri (Hasse) Vauterin et al., is one of the main problems affecting citrus production. In order to obtain resistance to phytopathogenic bacteria, insect genes, coding for antimicrobial proteins, have been used in plant genetic transformation. In this study, transgenic Citrus sinensis (L.) Osb. `Hamlin' plants expressing the antimicrobial insect-derived attacin A gene (attA) were obtained by Agrobacterium tumefaciens (Smith and Towns.) Conn-mediated transformation. Initially, the cDNA clone was used to construct a binary plasmid vector (pCattA 2300). The construction included the native signal peptide (SP) responsible for directing the insect protein to the extracellular space where bacteria is supposed to accumulate in vivo. In order to investigate the native SP effectiveness in a plant model system, onion (Allium cepa L.) epidermal cells were transformed, via biobalistics, using plasmids containing the attA gene with or without SP, fused with the green fluorescent protein gene (pattA 1303 and pSPattA 1303). Fluorescence accumulation surrounding the cells was observed only in tissues transformed with the plasmid containing the gene with SP, indicating the protein secretion to the apoplast. Citrus transformation was confirmed by PCR and Southern blot hybridization analysis in 12 regenerated plants. Transcription of attA gene was detected by Northern blot analysis in all transgenic plants. Eight selected transgenic lines were propagated and inoculated with a 106 cfu/mL suspension of the pathogen X. axonopodis pv. citri. Compared to control (non-transformed plant), seven transgenic lines showed a significant reduction in susceptibility to citrus canker. The results obtained here indicate the potential use of antibacterial proteins to protect citrus from bacterial diseases.

scholarly journals Improvement of a Genetic Transformation System and Preliminary Study on the Function of LpABCB21 and LpPILS7 Based on Somatic Embryogenesis in Lilium pumilum DC. Fisch

2020 ◽  
Vol 21 (18) ◽  
pp. 6784
Author(s):  
Shengli Song ◽  
Rui Yan ◽  
Chunxia Wang ◽  
Jinxia Wang ◽  
Hongmei Sun
Keyword(s):  
Somatic Embryogenesis ◽  
Genetic Transformation ◽  
Somatic Embryo ◽  
Asymmetric Distribution ◽  
Embryo Induction ◽  
Transgenic Lines ◽  
Preliminary Study ◽  
Genetic Transformation System ◽  
Mutant Lines ◽  
Somatic Embryo Induction

Auxin transport mediates the asymmetric distribution of auxin that determines the fate of cell development. Agrobacterium-mediated genetic transformation is an important technical means to study gene function. Our previous study showed that the expression levels of LpABCB21 and LpPILS7 are significantly up-regulated in the somatic embryogenesis (SE) of Lilium pumilum DC. Fisch. (L. pumilum), but the functions of both genes remain unclear. Here, the genetic transformation technology previously developed by our team based on the L.pumilum system was improved, and the genetic transformation efficiency increased by 5.7–13.0%. Use of overexpression and CRISPR/Cas9 technology produced three overexpression and seven mutant lines of LpABCB21, and seven overexpression and six mutant lines of LpPILS7. Analysis of the differences in somatic embryo induction of transgenic lines confirmed that LpABCB21 regulates the early formation of the somatic embryo; however, excessive expression level of LpABCB21 inhibits somatic embryo induction efficiency. LpPILS7 mainly regulates somatic embryo induction efficiency. This study provides a more efficient method of genetic transformation of L. pumilum. LpABCB21 and LpPILS7 are confirmed to have important regulatory roles in L. pumilum SE thus laying the foundation for subsequent studies of the molecular mechanism of Lilium SE.

GENETIC TRANSFORMATION OF APPLE WITH BIBAC VECTORS TO INCREASE SCAB RESISTANCE IN TRANSGENIC LINES

2007 ◽  
pp. 361-367
Author(s):  
J. Zhu ◽  
A. Zhou ◽  
S. Granozio ◽  
E. Andruccioli ◽  
E. Belfanti ◽  
...  
Keyword(s):  
Genetic Transformation ◽  
Scab Resistance ◽  
Transgenic Lines

Novel insights into the Citrus sinensis nonhost response suggest photosynthesis decline, abiotic stress networks and secondary metabolism modifications

2015 ◽  
Vol 42 (8) ◽  
pp. 758 ◽  
Author(s):  
Lucas D. Daurelio ◽  
M. Laura Tondo ◽  
M. Soledad Romero ◽  
Paz Merelo ◽  
Adriana A. Cortadi ◽  
...  
Keyword(s):  
Abiotic Stress ◽  
Secondary Metabolism ◽  
Citrus Sinensis ◽  
Xanthomonas Campestris ◽  
Carotenoid Biosynthesis ◽  
Citrus Canker ◽  
Stress Factors ◽  
Breeding Programs ◽  
Living Organisms ◽  
Shock Proteins

Plants are constantly exposed to stress factors. Biotic stress is produced by living organisms such as pathogens, whereas abiotic stress by unfavourable environmental conditions. In Citrus species, one of the most important fruit crops in the world, these stresses generate serious limitations in productivity. Through biochemical and transcriptomic assays, we had previously characterised the Citrus sinensis (L.) Osbeck nonhost response to Xanthomonas campestris pv. vesicatoria (Doidge), in contrast to Asiatic citrus canker infection caused by Xanthomonas citri subsp. citri (Hasse). A hypersensitive response (HR) including changes in the expression of several transcription factors was reported. Here, a new exhaustive analysis of the Citrus sinensis transcriptomes previously obtained was performed, allowing us to detect the over-representation of photosynthesis, abiotic stress and secondary metabolism processes during the nonhost HR. The broad downregulation of photosynthesis-related genes was correlated with an altered photosynthesis physiology. The high number of heat shock proteins and genes related to abiotic stress, including aquaporins, suggests that stresses crosstalk. Additionally, the secondary metabolism exhibited lignin and carotenoid biosynthesis modifications and expression changes in the cell rescue GSTs. In conclusion, novel features of the Citrus nonhost HR, an important part of the plants’ defence against disease that has yet to be fully exploited in plant breeding programs, are presented.

scholarly journals Comparison between Protoplast Transformation and Co-transformation in `Hamlin' Sweet Orange [Citrus sinensis (L.) Osbeck]

HortScience ◽  
2006 ◽  
Vol 41 (4) ◽  
pp. 972D-973
Author(s):  
Ahmad A. Omar ◽  
Wen-Yuan Song ◽  
James H. Graham ◽  
Jude W. Grosser
Keyword(s):  
Transgenic Plants ◽  
Citrus Sinensis ◽  
Fluorescent Protein ◽  
Sweet Orange ◽  
Citrus Canker ◽  
Yield Potential ◽  
High Yield ◽  
Pcr Analysis ◽  
Dna Encoding ◽  
Early Maturity

Citrus canker disease caused by the bacterial pathogen Xanthomonas axonopodis pv. citri is becoming a worldwide problem. Xa21 gene is a member of the Xa21 gene family of rice, which provides broad spectrum Xanthomonas resistance in rice. `Hamlin' sweet orange [Citrus sinensis (L.) Osbeck) is one of the leading commercial cultivars in Florida because of its high yield potential and early maturity. `Hamlin' also has a high regeneration capacity from protoplasts and is often used in transformation experiments. Since the citrus canker pathogen is in the same genus, this gene may have potential to function against canker in citrus. The wild-type Xa21 gene contains an intron, and there are some questions whether dicot plants can process genes containing monocot introns (the cDNA is intron-free). Plasmids DNA, encoding the non-destructive selectable marker EGFP (Enhanced Green Fluorescent Protein) gene and the cDNA of the Xa21 gene were transformed or co-transformed into `Hamlin' orange protoplasts using polyethylene glycol. More than 200 transgenic embryoids were recovered. More than 400 transgenic plants were developed from 75 independent transgenic events. PCR analysis revealed the presence of the cDNA of the Xa21 and the GFP genes in the transgenic plants. Some of the plants have the GFP only. Southern analysis is showing integration of the cDNA into different sites ranges from one to five sites. Western analysis is showing the expression of the cDNA of the Xa21 gene in the transgenic citrus plants. This is the first time that a gene from rice has been stably integrated and expressed in citrus plants. Canker challenge assay is in progress.

Transformation of sweet orange [Citrus sinensis (L.) Osbeck] with pthA-nls for acquiring resistance to citrus canker disease

2010 ◽  
Vol 75 (1-2) ◽  
pp. 11-23 ◽  
Author(s):  
Li Yang ◽  
Chunhua Hu ◽  
Na Li ◽  
Jiayin Zhang ◽  
Jiawen Yan ◽  
...  
Keyword(s):  
Citrus Sinensis ◽  
Sweet Orange ◽  
Citrus Canker ◽  
Canker Disease ◽  
Citrus Canker Disease

scholarly journals Agrobacterium-mediated transformation of Citrus sinensis and Citrus limonia epicotyl segments

2003 ◽  
Vol 60 (1) ◽  
pp. 23-29 ◽  
Author(s):  
Weliton Antonio Bastos de Almeida ◽  
Francisco de Assis Alves Mourão Filho ◽  
Beatriz Madalena Januzzi Mendes ◽  
Alexandra Pavan ◽  
Adriana Pinheiro Martinelli Rodriguez
Keyword(s):  
Genetic Transformation ◽  
Citrus Sinensis ◽  
Transformation Efficiency ◽  
Petri Dish ◽  
Cultivation Medium ◽  
Breeding Programs ◽  
Rangpur Lime ◽  
Citrus Breeding ◽  
In Vitro Plantlets

Genetic transformation allows the release of improved cultivars with desirable characteristics in a shorter period of time and therefore may be useful in citrus breeding programs. The objective of this research was to establish a protocol for genetic transformation of Valencia and Natal sweet oranges (Citrus sinensis L. Osbeck) and Rangpur lime (Citrus limonia L. Osbeck). Epicotyl segments of germinated in vitro plantlets (three weeks in darkness and two weeks in a 16-h photoperiod) were used as explants. These were co-cultivated with Agrobacterium tumefaciens strain EHA-105 and different experiments were done to evaluate the transformation efficiency: explants were co-cultivated with Agrobacterium for one, three or five days; explants were incubated with Agrobacterium suspension for 5, 10, 20 or 40 minutes; co-cultivation medium was supplemented with acetosyringone at 0, 100 or 200 µmol L-1; Explants ends had a longitudinal terminal incision (2-3 mm); co-cultivation temperatures of 19, 23 or 27°C were imposed. The experimental design was completely randomized in all experiments with five replications, each consisted of a Petri dish (100 x 15 mm) with 30 explants and resulted in a total of 150 explants per treatment. Longitudinal terminal incision in the explant ends did not improve shoot regeneration. However, transgenic plants of all three cultivars were confirmed from explants that had been subjected to inoculation time of 20 minutes, co-culture of three days at 23-27°C, in the absence of acetosyringone.

Sign in / Sign up

Export Citation Format

Share Document