May 07, 2026
An efficient regeneration and transformation protocol for the potato cv. Kufri Jyoti
- Vandna Patial1,
- Manavi Raizada2,
- Ramawatar Nagar2
- 1ICAR-Indian Agricultural Research Institute;
- 2ICAR-National Institute for Plant Biotechnology
- Vandna Patial: The conception, design, or validation of the protocol;
- Manavi Raizada: Data acquisition relevant to the protocol
- Plant transformation and genome editing
Protocol Citation: Vandna Patial, Manavi Raizada, Ramawatar Nagar 2026. An efficient regeneration and transformation protocol for the potato cv. Kufri Jyoti. protocols.io https://dx.doi.org/10.17504/protocols.io.rm7vz4pm8lx1/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: In development
We are still developing and optimizing this protocol
Created: April 24, 2026
Last Modified: May 07, 2026
Protocol Integer ID: 315650
Keywords: potato, kurfi jyoti, regeneration, transformation, Solanum tuberosum, regeneration system for potato cv, transformation protocol for the potato cv, major challenge for crop improvement, plant genetic transformation strategy, crop improvement, resilient crop variety, potato starch, recalcitrant nature of many elite crop genotype, potato cv, many elite crop genotype, important food crop for human consumption, important food crop, efficient producers of edible energy, potato, optimised regeneration system, efficient regeneration, genetic transformation strategy, powerful tools for genetic engineering, rice, genetic engineering, regeneration, edible energy, efficient producer, rich source of carbohydrate, wheat, advanced molecular breeding, important component in the production, kufri jyoti, carbohydrate, global food security
Abstract
Beyond being the third most important food crop for human consumption after rice and wheat, the potato is also one of the most efficient producers of edible energy per unit area of land. As a nutrient-dense food, potatoes are a rich source of carbohydrates, vitamins, and minerals. Potato starch is equally valued for its functional properties in industrial applications, serving as an important component in the production of chemical additives and as a sizing agent in the textile and paper industries.
Climate change and rapid population growth necessitate the development of high-yielding, nutritionally enhanced, and climate-resilient crop varieties to ensure global food security. Plant genetic transformation strategies provide powerful tools for genetic engineering and advanced molecular breeding; however, the recalcitrant nature of many elite crop genotypes remains a major challenge for crop improvement. Therefore, the present study describes an optimised regeneration system for potato cv. Kufri Jyoti.
Materials
Biological Materials: Nodal explants from ex-vitro grown potato seedlings.
Chemicals: double distilled water, triton X, 1% bavistein, sodium hypochlorite (4%) solution, 70 % ethanol, Murashige and Skoog medium, NAA, zeatin, GA3, sucrose, agar
Glasswares: test tubes, jam bottles, conical flasks, beakers,
Other Materials: forceps, scalpel, scissors, petriplates, test tube stand, filter paper, cotton, blotting sheet, parafilm, burner/ lamp, matchbox, micropipette, and its tips, soilrite, pots, clear plastic bags
Before start
- Take healthy internodal cuttings as starting material.
- The pH of all media used was adjusted to 5.6- 5.8.
- Media was sterilized using autoclaving at 121°C and 105 kPa for 20 minutes.
- All the instruments used e.g. forceps, scalpel, should be properly sterilized.
Surface sterilization of Potato nodes
35m
Potato nodes were immersed in water having few drops of tween 20 for 5-8 minutes. This step facilitates removal of surface impurities. followed by thoroughly rinsing in running tap water for 10-15 minutes.
20m
Then explants were transferred to laminar airflow cabinet for further sterilization using 70% ethanol (v/v) for 1-2 minutes.
2m
This was followed by 7-8 minutes of 4% sodium hypochlorite (v/v), to eliminate any surface microorganisms.
8m
Finally, explants underwent several rinses (3-5) with autoclaved doubled distilled water before inoculation, which ensured the removal of any residual ethanol and sodium hypochlorite.
Note:
Nodes should be sterilized properly to prevent any contamination.
5m
Inoculation
After sterilization, nodes were inoculated on hormone-free Murashige and Skoog (MS) medium supplemented with 3% sucrose and 0.8% agar.
Note:
The cultures were maintained in a tissue culture room with a light intensity of 2000 lux, temperature control at 24 ± 2 °C and a photoperiod of 16 h light/8 h dark.
Shoot Regeneration
The internodal stem (1-2cm) cuttings from in-vitro raised plants were used as explants for regeneration and inoculated on regeneration medium.
Shoot regeneration started after 6-7 days of inoculation. The internodal stem cuttings (1-2cm) from in-vitro raised aseptic plants were used as explants for regeneration experiments and inoculated on regeneration medium.
Regeneration medium: MS+ NAA (1 mg/l) + GA3 (2 mg/l) + zeatin (1 mg/l) + sucrose (3%) + agar (0.8%)
Note:
The cultures were kept at 24±2°C with 16h of light and 8h of dark photoperiod. All the cultures were sub-cultured at 15 days interval.
Rhizogenesis
For rhizogenesis, 3-4 cm long regenerated shoots were transferred to rooting medium
Rhizogenesis medium: MS+ sucrose (3%) + agar (0.8%)
Acclimatization
After 15-20 days, well-rooted plantlets were gently removed from medium using long forceps.
Roots were washed with lukewarm water to clear any tissue culture medium left.
In-vitro regenerated plantlets were transplanted to pots containing sterilized soilrite and covered with clear plastic bags to maintain high humidity while the plants become established in soil. These plantlets were placed in the growth chamber at 23±2 °C with 16 hrs. photoperiod.
After 1 week, plastic bags were removed. Hardened plants were grown under standard green house conditions until maturity. On maturity, micro tubers were harvested.
Note:
Care should be taken while removing the in vitro grown plantlets from medium.
Regeneration and transformation of potato variety Kufri Jyoti. a and b: Inoculation of in vitro internodal segments on MS + NAA + Zeatin + GA₃ supplemented medium. c: callusing after 2 weeks of inoculation. d: Development of shoots in putative transformants. e: Rooting of regenerated shoots. f: full grown plants in pots