Introduction: Anther culture is a type of tissue culture technique, where a part of a plant, i.e. Anther gives rise to an androgenic haploid plant when cultured in a nutrient medium under optimal conditions that are required for the growth of the cell. The process of anther culture comes under the process of “Androgenesis” which merely refers to the induction of haploid plants.
In about 250 species, the technique of the anther culture has been implemented. Among these 250 species, the most common species belongs to the family of Gramineae, Solanaceae, Ranunculaceae, Cruciferae etc.
Content: Anther Culture
- What is Anther?
- History of Anther Culture
- Definition of Anther Culture
- Objective of Anther Culture
- Methods in Anther Culture
- Protocol of Anther Culture
- Factors influencing Anther Culture
What is Anther?
The anther is a part of a flower which is bilobed and sac-like in structure. In flower, anther is the part of stamen which is a male reproductive part. Stamen comprises a long, narrow or stalk-like filament which carries the anther at its tip.
An anther consists of microsporangia which produce pollen grains by the meiotic cell division.
History of Anther Culture
There are several discoveries that have led to the discovery of producing haploid plantlets by the anther culture. Many scientists have contributed to let us know more about this technique by their experiments.
|Year of discovery||Discovered by||Name of the discovery|
|1921||Bergner||Introduced the existence of haploids in Datura stramonium|
|1953||W.Tulecke||Introduced the proliferation of haploid callus from the mature pollen grains of Ginkgo biloba|
|1966||Guha and S.C. Maheshwar||Confirmed the origin of haploid plantlets from the anther|
|1967||Bourgin and Hitsch||Discovered the full-fledged haploid plants from the Nicotiana tabacum.|
Definition of Anther Culture
Anther culture is a method of Androgenesis, i.e. production of haploid plantlets by the in-vitro condition where the anther is separated from the bud and is followed by sterilization and culturing in nutrient media to give rise to the haploid plantlets.
Objective of Anther Culture
The primary objective of the anther culture technique is to produce haploid androgenic plants by using the regeneration capacity or the totipotency of the anther.
Methods in Anther Culture
Anther culture is a plant tissue culture method which can be done by two methods either through a direct or indirect method.
- The direct method of an anther culture involves “Embryogenesis”. In this method, the anther behaves as a “Zygote” and forms embryoid which gives rise to haploid plantlet.
- The indirect method of an anther culture involves “Organogenesis”. In this method, the anther undergoes cell division repeatedly to form the callus tissue which later gives rise to the haploid plantlets.
Protocol of Anther Culture
There are following steps involved in the anther culture method which includes:
- First, select the unopened buds of size about 17-22mm in length. Reject the buds that are beginning to open or already open. While selecting buds, the size of the sepal is equal to the size of a petal is ideal for the anther culture.
- Transfer the buds into sterilized airtight plastic bags.
- Then, transfer the selected buds to the laminar airflow chamber.
- After that surface sterilization of buds is carried out in the chamber to maintain the aseptic conditions. Firstly, the buds are surface sterilized by 70% ethanol for 10seconds and then 20% of sodium hypochlorite for 10minutes.
- Then wash the buds three times by distilled water.
- After washing, transfer the buds to the sterilized Petri plate.
- Then with the help of a scalpel, separate the stamen from the bud.
- Remove the filaments from an anther.
- After that, transfer an anther onto the solid or liquid nutrient medium and incubate it for 3-4 weeks at 24-28 ֯C in the dark.
- Then either by embryogenesis and organogenesis, haploid plantlets would appear from the anther culture 3-4 weeks. During this stage, incubate the culture at 24-28 ֯C for 12-18 hours in light and for 6-12 hours in the dark.
- At last, about 50mm tall plantlets appear which then transfer into the pot containing bio compost followed by washing. Then a sterilized glass beaker is used to cover the pot and remove the glass beaker after some week and transfer the plant to the large pot.
Factors influencing anther culture
There are some factors influences the technique of the anther culture includes some factors which can categorize into intrinsic and extrinsic factors.
Anther wall: It provides nourishment in the developmental stages of the anther.
Stage of Anther: For anther culture pre-mitotic, mitotic and post-mitotic stages of an anther is prefer. Pre-mitotic is the stage, when the first meiotic division occurs in the anther and where the pollens are at an immature stage. In the mitotic stage of an anther, there is the division of the pollen. The post-mitotic stage is the bi-cellular stage where development in pollen grains occurs to form embryoids.
Bud size: The anther culture prefers the size of bud that is up to 17-22mm in length.
Age of plant: The anther culture prefers the buds from the younger plants.
Culture medium: In the culture medium sucrose, iron, vitamins, coconut milk, hormones and growth regulators (Auxin or cytokinin) plays an essential role in the induction of haploid plant.
Temperature: It varies with the different plant species.
Example: In Datura stramonium, the optimum temperature is 20 ֯ C for the formation of embryoids. In Nicotiana tabacum, the optimum temperature is 25 ֯ C for the formation of embryoids. In Brassica campentris, the optimum temperature is much higher that is 35 ֯ C for the formation of embryoids.
Role of activated charcoal: Activated charcoal plays a vital role in the removal of inhibitory substances like both exo and endogenous growth hormones from the culture medium by stimulating the growth of an anther.
Pre-treatment method: For the anther culture, pre-treatment of the bud is also suitable for the production of embryoids.
Example: In Nicotiana tabacum, the bud is pretreated at a temperature of 5 ֯ C for 72 hours.