Somatic embryogenesis and Agrobacterium-mediated transformation of Gladiolus hybridus cv. ‘Advance Red’

Wu, Jian; Liu, Chen; Seng, Shanshan; Khan, Muhammad Ali; Sui, Juanjuan; Bi, Gong; Liu, Chao; Wu, Chenyu; Zhong, X.P.; He, Junna; Yi, Mingfang

doi:10.1007/s11240-014-0639-5

Cited by 26 publications

(11 citation statements)

References 60 publications

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“…In addition, although glutamine or hydrolyzed casein alone had no significant effect on the embryo yield, their combined effect on somatic embryogenesis was significant. The major factors affecting Agrobacteriummediated transformation of conifers are the physiological status of the plant recipient, the bacterial strain and plasmid vectors, and the infection and screening conditions (Wu et al, 2015;Kim et al, 2016;Aggarwal et al, 2018;Li et al, 2018). Furthermore, Agrobacterium load and the infection time are also important determinants for successful transformation (Li et al, 2017(Li et al, , 2018.…”

Section: Discussionmentioning

confidence: 99%

Stable and Efficient Agrobacterium-Mediated Genetic Transformation of Larch Using Embryogenic Callus

et al. 2020

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Larix olgensis or larch is an economically important coniferous tree species with rapid growth in the early stages, strong adaptability, and a short time to harvest. The genetic improvement of larch has garnered considerable attention in recent years for reclaiming timber forests. However, traditional breeding methods are largely ineffective for achieving rapid genetic improvement of L. olgensis. Studies show that the efficiency of plant regeneration can be improved by optimizing somatic embryogenesis. On this basis, we devised a stable, fast and efficient Agrobacterium-mediated genetic transformation method using suspended embryogenic calluses as explants and β-glucuronidase as the reporter. We evaluated the effects of the Agrobacterium load, co-culture period, and addition of acetosyringone and transformant screening antibiotic on the transformation efficiency. In addition, we tested the pCAMBIA 1300-PtHCA 2-1 promoter-GUS binary expression vector, which contains the GUS gene ORF under the control of Populus trichocarpa high cambial activity PtHCA 2-1 promoter, and observed the tissue-specific expression of the GUS gene in the somatic embryos of transgenic larch. This novel technique can not only accelerate the generation of superior transgenic strains of L. olgensis but also aid in future gene functional studies.

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Section: Discussionmentioning

confidence: 99%

Stable and Efficient Agrobacterium-Mediated Genetic Transformation of Larch Using Embryogenic Callus

et al. 2020

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“…SE has been reported in Arabidopsis thaliana (Gaj, 2001), Phoenix dactylifera L. (Naik and Al-Khayri, 2016), Theobroma cacao L. (Ajijah et al, 2016), Gladiolus hybridus (Wu J. et al, 2015), and Cassia occidentalis L. (Naz et al, 2015). However, the understanding of the mechanism underlying the formation of somatic embryos is still in its infancy, and SE remains the least understood pattern of regeneration (Fehér, 2015).…”

Section: Introductionmentioning

confidence: 97%

Small RNA and Transcriptome Sequencing Reveal a Potential miRNA-Mediated Interaction Network That Functions during Somatic Embryogenesis in Lilium pumilum DC. Fisch.

et al. 2017

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Plant somatic embryos are widely used in the fields of germplasm conservation, breeding for genetic engineering and artificial seed production. MicroRNAs (miRNAs) play pivotal roles in somatic embryogenesis (SE) regulation. However, their regulatory roles during various stages of SE remain unclear. In this study, six types of embryogenic samples of Lilium pumilum DC. Fisch., including organogenic callus, embryogenic callus induced for 4 weeks, embryogenic callus induced for 6 weeks, globular embryos, torpedo embryos and cotyledon embryos, were prepared for small RNA sequencing. The results revealed a total of 2,378,760 small RNA reads, among which the most common size was 24 nt. Four hundred and fifty-two known miRNAs, belonging to more than 86 families, 57 novel miRNAs and 40 miRNA*s were identified. The 86 known miRNA families were sorted according to an alignment with their homologs across 24 land plants into the following four categories: 23 highly conserved, 4 moderately conserved, 15 less conserved and 44 species-specific miRNAs. Differentially expressed known miRNAs were identified during various stages of SE. Subsequently, the expression levels of 12 differentially expressed miRNAs and 4 targets were validated using qRT-PCR. In addition, six samples were mixed in equal amounts for transcript sequencing, and the sequencing data were used as transcripts for miRNA target prediction. A total of 66,422 unigenes with an average length of 800 bp were assembled from 56,258,974 raw reads. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment indicated that 38,004 and 15,497 unigenes were successfully assigned to GO terms and KEGG pathways, respectively. Among the unigenes, 2,182 transcripts were predicted to be targets for 396 known miRNAs. The potential targets of the identified miRNAs were mostly classified into the following GO terms: cell, binding and metabolic process. Enriched KEGG analysis demonstrated that carbohydrate metabolism was the predominant pathway in Lilium SE. Thus, we performed systemic characterization, homology comparisons and profiling of miRNA expression, and we constructed an miRNA-target network during Lilium SE for the first time. Our findings establish a foundation for the further exploration of critical genes and elucidation of SE in Lilium.

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“…Transformation conditions are key factors affecting genetic transformation. It is generally believed that the pre-culture of explants before transformation can promote cell division, make it easier to integrate exogenous genes, and improve the level of transient expression and transformation rate of exogenous genes [52,53]. Different explants have different pre-culture durations.…”

Section: Discussionmentioning

confidence: 99%

“…Co-culture durations and gene transfer and integration into the plant genome via Agrobacterium vary widely, e.g., from several hours to several days, depending on plant species, explant type and culture conditions [52,56]. It has been reported that co-culture has significant effects on the number of GUS-positive calli in rice (3 days) [52], Lycopersicon esculentum (3–4 days) [57], Rosa chinensis (2 days) [58], Epipremnum aureum (5 days) [59] and Gladiolus hybridus (12 days) [53]. In this study, co-culture for 3 days had the best effect, and transformation efficiency could not be improved by prolonging co-culture duration in Lilium transformation.…”

Section: Discussionmentioning

confidence: 99%

Establishment of Efficient Genetic Transformation Systems and Application of CRISPR/Cas9 Genome Editing Technology in Lilium pumilum DC. Fisch. and Lilium longiflorum White Heaven

Yan

Wang

Ren

et al. 2019

IJMS

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Lilium spp. is a bulb flower with worldwide distribution and unique underground organs. The lack of an efficient genetic transformation system for Lilium has been an international obstacle. Because existing model plants lack bulbs, bulb-related gene function verification studies cannot be carried out in model plants. Here, two stable and efficient genetic transformation systems based on somatic embryogenesis and adventitious bud regeneration were established in two Lilium species. Transgenic plants and T-DNA insertion lines were confirmed by β-glucuronidase (GUS) assay, polymerase chain reaction (PCR) and Southern blot. After condition optimization, transformation efficiencies were increased to 29.17% and 4% in Lilium pumilum DC. Fisch. and the Lilium longiflorum ‘White Heaven’, respectively. To further verify the validity of these transformation systems and apply the CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated protein 9) technology in Lilium, the LpPDS gene in the two Lilium species was knocked out. Completely albino, pale yellow and albino–green chimeric mutants were observed. Sequence analysis in the transgenic lines revealed various mutation patterns, including base insertion, deletion and substitution. These results verified the feasibility and high efficiency of both transformation systems and the successful application of the CRISPR/Cas9 system to gene editing in Lilium for the first time. Overall, this study lays an important foundation for gene function research and germplasm improvement in Lilium spp.

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Somatic embryogenesis and Agrobacterium-mediated transformation of Gladiolus hybridus cv. ‘Advance Red’

Cited by 26 publications

References 60 publications

Stable and Efficient Agrobacterium-Mediated Genetic Transformation of Larch Using Embryogenic Callus

Stable and Efficient Agrobacterium-Mediated Genetic Transformation of Larch Using Embryogenic Callus

Small RNA and Transcriptome Sequencing Reveal a Potential miRNA-Mediated Interaction Network That Functions during Somatic Embryogenesis in Lilium pumilum DC. Fisch.

Establishment of Efficient Genetic Transformation Systems and Application of CRISPR/Cas9 Genome Editing Technology in Lilium pumilum DC. Fisch. and Lilium longiflorum White Heaven

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