scholarly journals In Planta Transformation in Plants: A Review

2019 ◽  
Vol 40 (03) ◽  
Author(s):  
Ratna Preeti Kaur ◽  
Sugani Devi
Keyword(s):  
Tissue Culture ◽  
Developmental Stages ◽  
Crop Improvement ◽  
Induction Medium ◽  
Foreign Gene ◽  
In Planta Transformation ◽  
In Planta ◽  
Agrobacterium Strain ◽  
Plant Parts ◽  
Whole Plants

In planta transformation has been established as an innovative and simple technique involving direct transformation of plant parts without involving the tedious tissue culture step. Methodologies of in planta transformation involve different plant parts and strategies. Agrobacterium strain carrying the gene of interest is targeted to the specific plant part either directly or in the induction medium. Vacuum infiltration is sometimes used to facilitate foreign gene integration. Initial studies of this novel technique involved treatment of whole plants with the inoculum, and later shifted to treatments of shoot tips and floral parts and finally the female reproductive parts have been targeted. Zygotes, embryos and seeds have recently been used extensively yielding successful transformation. The method is simple, convenient and overcomes the problem of tissue culture induced genetic variability in the transformants. The review traces the origin and development in the in planta methodologies used over the past and the various parameters considered by the workers for increased effectiveness viz. developmental stages, Agrobacterium stain, surfactant, induction medium etc., in the various crops compiled. Based on the review it may be inferred that there is immense potential in planta transformation, and the ease of regeneration and selection of transformants in the methods described, can be utilized for crop improvement.

scholarly journals Tissue culture-based Agrobacterium-mediated and in planta transformation methods

2017 ◽  
Vol 53 (No. 4) ◽  
pp. 133-143 ◽  
Author(s):  
M. Niazian ◽  
S.A. Sadat Noori ◽  
P. Galuszka ◽  
S.M.M. Mortazavian
Keyword(s):  
Tissue Culture ◽  
Transformation Method ◽  
Gene Transformation ◽  
In Planta Transformation ◽  
In Planta ◽  
Agrobacterium Mediated Transformation ◽  
Advantages And Disadvantages ◽  
Culture Independent ◽  
Transformation Methods ◽  
Mutant Lines

Gene transformation can be done in direct and indirect (Agrobacterium-mediated) ways. The most efficient method of gene transformation to date is Agrobacterium-mediated method. The main problem of Agrobacterium-method is that some plant species and mutant lines are recalcitrant to regeneration. Requirements for sterile conditions for plant regeneration are another problem of Agrobacterium-mediated transformation. Development of genotype-independent gene transformation method is of great interest in many plants. Some tissue culture-independent Agrobacterium-mediated gene transformation methods are reported in individual plants and crops. Generally, these methods are called in planta gene transformation. In planta transformation methods are free from somaclonal variation and easier, quicker, and simpler than tissue culture-based transformation methods. Vacuum infiltration, injection of Agrobacterium culture to plant tissues, pollen-tube pathway, floral dip and floral spray are the main methods of in planta transformation. Each of these methods has its own advantages and disadvantages. Simplicity and reliability are the primary reasons for the popularity of the in planta methods. These methods are much quicker than regular tissue culture-based Agrobacterium-mediated gene transformation and success can be achieved by non-experts. In the present review, we highlight all methods of in planta transformation comparing them with regular tissue culture-based Agrobacterium-mediated transformation methods and then recently successful transformations using these methods are presented.

scholarly journals CURRENT STATUS OF TRANSGENIC COTTON-UTILITY OF GENOTYPE INDEPENDENT IN PLANTA APPROACHES FOR THE GENERATION OF TRANSGENICS

2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Kesiraju Karthik
Keyword(s):  
Tissue Culture ◽  
In Vitro Regeneration ◽  
Crop Improvement ◽  
Transgenic Cotton ◽  
Current Status ◽  
In Planta ◽  
Biotic And Abiotic Stresses ◽  
Independent Manner ◽  
Gossypium Spp

Cotton (Gossypium spp.), is a mercantile crop plant is grown for its fluffy fiber and cotton seed oil in around 70 countries worldwide. Cotton is an economically important crop, shows erratic productivity under rain feed conditions; it is bogged down with many biotic and abiotic stresses. Due to lack of resistant germplasm, crop improvement through conventional breeding practices has been lagging. Genetic engineering offers numerous protocols to engineer plants to overcome stress. Biotechnological intervention for cotton improvement has begun three decades ago. The recalcitrance of cotton to tissue culture has been the major constraint for in vitro regeneration. Alternate methods that evade tissue culture regeneration steps have thus been envisaged. Till date there are very few standardized protocols that can be employed to develop transgenics in a genotype independent manner. Thus, genotype independent in planta transformation strategies have gained momentum in the present days, but reproducibility of reported protocols remains an amigna in many cases. In planta transformations holds prominence due to viability and ease in generation of transgenic cotton plants with in less time. This review focuses on grouping efforts made by different research groups in this senior. Several reports and standardizations have been focused that reports development of transgenic cotton.

scholarly journals Agrobacterium tumefaciens-MEDIATED IN PLANTA TRANSFORMATION METHOD FOR THE SoSPS1 GENE IN CITRUS PLANTS (Citrus nobilis L.)

2020 ◽  
Vol 7 (1) ◽  
pp. 31
Author(s):  
Ni Putu Ayu Erninda Oktaviani Suputri ◽  
Rindang Dwiyani ◽  
Ida Ayu Putri Darmawanti ◽  
Bambang Sugiharto
Keyword(s):  
Tissue Culture ◽  
Agrobacterium Tumefaciens ◽  
Cauliflower Mosaic Virus ◽  
Total Soluble Protein ◽  
35S Promoter ◽  
Nptii Gene ◽  
Main Function ◽  
In Planta Transformation ◽  
In Planta ◽  
Regeneration System

The SoSPS1 gene of sugar cane plants previously subjected to Agrobacterium tumefacienmediated cloning was to be transferred to citrus plants to increase metabolism of sucrose in plant. The T-DNA harbored the SoSPS1 gene under the control of the CaMV 35S promoter from the cauliflower mosaic virus and contained the NPTII gene (kanamycin resistance gene) as a selectable marker for transformant selection. Generally, gene transformation in plants is carried out by tissue culture. However, tissue culture has several disadvantages such as its being time-consuming, its sometimes resulting in somatic mutations and somaclonal variations, and the requirement of sterile conditions in the procedure of gene transfer. In planta transformation is a useful system for those plants that lack tissue culture and regeneration system. The main function of in planta transformation is to recover the advantages of tissue culture as an efficient, quick method, including its ability to produce a large number of transgenic plants and to accumulate a high concentration of total soluble protein in short time. There are two procedures of in planta transformation for the seeds of citrus plants, namely “prick and coat” and “seed tip-cutting and imbibition”. In the prick and coat method, seeds are pricked on their entire surfaces and smeared with a suspension of Agrobacterium tumefaciens. In the seed tip-cutting and imbibition method, on the other hand, seeds are cut at the tip and soaked in a suspension of Agrobacterium tumefaciens. The leaves derived from seeds treatment were taken as samples for DNA extraction and PCR using primers of the NPTII gene (Forward: 5’-GTCATCTCACCTTCCTCCTGCC-3’; Reverse: 5’-GTCGCTTGGTCGGTCATTTCG-3’). This research found that only the seed tip-cutting and imbibition plants amplified along the 550-bp band, while those of the prick and coat method did not. Additionally, the T-DNA was successfully integrated into the genome of the plants treated with the seed tip-cutting and imbibition method but not with the prick and coat.

scholarly journals PRICK AND SOAK Agroacterium tumefaciens-MEDIATED IN PLANTA TRANSFORMATION IN TOMATO (Lycopersicon esculentum Mill.)

2018 ◽  
Vol 5 (2) ◽  
pp. 124
Author(s):  
I Putu Wahyu Sanjaya ◽  
Rindang Dwiyani ◽  
I Gede Putu Wirawan ◽  
Bambang Sugiharto
Keyword(s):  
Tissue Culture ◽  
Agrobacterium Tumefaciens ◽  
Saccharum Officinarum ◽  
Plant Genome ◽  
Nptii Gene ◽  
In Planta Transformation ◽  
In Planta ◽  
Tomato Seeds ◽  
Inoculation Time

One of the modern plant breedings through genetic engineering is Agrobacterium tumefaciens-mediated transformation. Agrobacterium tumefaciens-mediated transformation can be performed in vitro or in planta. In planta transformation arises from the weaknesses of the in vitro method such as need high hygiene standard, professional tissue culture experts, and more time to prepare explants and somaclonal variation. In planta transformation is a method to transfer the gene to the plant genome without any tissue culture stages. The aims of this research were to know the possibility of the prick and soak in planta method with the target of tomato seeds and to know the most suitable inoculation time for tomato seeds transformation by prick and soak method the transformation is done by pricking the seeds and soaking them in the A. tumefaciens suspension. The treatments in this study were 1 and 2 days inoculation time to test the efficacy of prick and soak in planta transformation method. Tomato seeds were pricked with a needle on the center once, and then soaked in A. tumefaciens strain LB4404 suspension carrying pKYS-SoSPS1 plasmid with Neomycin Phosphotransferase (NPTII) and Saccharum officinarum Sucrose Phosphate synthase (SoSPS1) genes. Visualization of tomato’s DNA samples after PCR showed that 1-day inoculation sample was positively integrated with NPTII gene and negative in the 2 days inoculation treatment.

scholarly journals Ovule Development and in Planta Transformation of Paphiopedilum Maudiae by Agrobacterium-Mediated Ovary-Injection

2020 ◽  
Vol 22 (1) ◽  
pp. 84
Author(s):  
Bai-Xue Luo ◽  
Li Zhang ◽  
Feng Zheng ◽  
Kun-Lin Wu ◽  
Lin Li ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Developmental Stages ◽  
Embryo Sac ◽  
Mature Embryo ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Pcr Analysis ◽  
In Planta Transformation ◽  
In Planta ◽  
Hygromycin Selection

In this paper, the development of the Paphiopedilum Maudiae embryo sac at different developmental stages after pollination was assessed by confocal laser scanning microscopy. The mature seeds of P. Maudiae consisted of an exopleura and a spherical embryo, but without an endosperm, while the inner integument cells were absorbed by the developing embryo. The P. Maudiae embryo sac exhibited an Allium type of development. The time taken for the embryo to develop to a mature sac was 45-50 days after pollination (DAP) and most mature embryo sacs had completed fertilization and formed zygotes by about 50–54 DAP. In planta transformation was achieved by injection of the ovaries by Agrobacterium, resulting in 38 protocorms or seedlings after several rounds of hygromycin selection, corresponding to 2, 7, 5, 1, 3, 4, 9, and 7 plantlets from Agrobacterium-mediated ovary-injection at 30, 35, 42, 43, 45, 48, 50, and 53 DAP, respectively. Transformation efficiency was highest at 50 DAP (2.54%), followed by 2.48% at 53 DAP and 2.45% at 48 DAP. Four randomly selected hygromycin-resistant plants were GUS-positive after PCR analysis. Semi-quantitative PCR and quantitative real-time PCR analysis revealed the expression of the hpt gene in the leaves of eight hygromycin-resistant seedlings following Agrobacterium-mediated ovary-injection at 30, 35, 42, 43, 45, 48, 50, and 53 DAP, while hpt expression was not detected in the control. The best time to inject P. Maudiae ovaries in planta with Agrobacterium is 48-53 DAP, which corresponds to the period of fertilization. This protocol represents the first genetic transformation protocol for any Paphiopedilum species and will allow for expanded molecular breeding programs to introduce useful and interesting genes that can expand its ornamental and horticulturally important characteristics.

Australian Journal of Plant Physiology: Editorial report

1998 ◽  
Vol 25 (1) ◽  
pp. I
Author(s):  
Laurie Martinelli
Keyword(s):  
Plant Physiology ◽  
Linkage Analysis ◽  
Gene Tagging ◽  
Australian Journal ◽  
In Planta Transformation ◽  
In Planta ◽  
Novel Genes ◽  
Late Flowering ◽  
Allelism Tests ◽  
Whole Plants

Gene tagging with insertional mutagens greatly facilitates the isolation of novel genes. A new collection of Arabidopsis T-DNA tag insertion lines (n=2165) was generated by in planta transformation. Whole plants were vacuum-infiltrated in a suspension of Agrobacterium carrying the pGKB5 tagging vector. The efficiency of transformation increased with addition of the surfactant Silwet L-77 (0.005% v/v) to the Agrobacterium suspension. Visual screens of the T-DNA lines identified two mutants with floral defects. Allelism tests suggested that a mutation in the GIGANTEA gene was responsible for the late-flowering phenotype of one of the mutants. Linkage analysis indicated that the GIGANTEA gene was tagged in this mutant.

T-DNA tagging of a flowering-time gene and improved gene transfer by in planta transformation of Arabidopsis

1998 ◽  
Vol 25 (1) ◽  
pp. 125 ◽  
Author(s):  
Kim Richardson ◽  
Sarah Fowler ◽  
Carly Pullen ◽  
Caryl Skelton ◽  
Bret Morris ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Linkage Analysis ◽  
Flowering Time ◽  
Gene Tagging ◽  
In Planta Transformation ◽  
In Planta ◽  
Late Flowering ◽  
Allelism Tests ◽  
Flowering Time Gene ◽  
Whole Plants

Gene tagging with insertional mutagens greatly facilitates the isolation of novel genes. A new collection of Arabidopsis T-DNA tag insertion lines (n=2165) was generated by in planta transformation. Whole plants were vacuum-infiltrated in a suspension of Agrobacterium carrying the pGKB5 tagging vector. The efficiency of transformation increased with addition of the surfactant Silwet L-77 (0.005% v/v) to the Agrobacterium suspension. Visual screens of the T-DNA lines identified two mutants with floral defects. Allelism tests suggested that a mutation in the GIGANTEA gene was responsible for the late-flowering phenotype of one of the mutants. Linkage analysis indicated that the GIGANTEA gene was tagged in this mutant.

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