Key Applications of Plant Tissue Culture in Genetics and Breeding

Germplasm Conservation and Cryopreservation

In vitro cell and organ culture offers an alternative source for the conservation of endangered genotypes [40]. Germplasm conservation worldwide is increasingly becoming an essential activity due to the high rate of disappearance of plant species and the increased need for safeguarding the floristic patrimony of countries [41].

Tissue culture protocols can be used for preservation of vegetative tissues when the targets for conservation are clones instead of seeds, to keep the genetic background of a crop, and to avoid the loss of the conserved patrimony due to natural disasters, whether biotic or abiotic stress [42]. Plant species which do not produce seeds (sterile plants) or which have ‘recalcitrant’ seeds that cannot be stored for long periods of time can successfully be preserved via in vitro techniques for the maintenance of gene banks.

The Role of Cryopreservation

Cryopreservation plays a vital role in the long-term in vitro conservation of essential biological material and genetic resources. It involves the storage of in vitro cells or tissues in liquid nitrogen. This process, however, can result in cryo-injury upon the exposure of tissues to physical and chemical stresses.

Successful cryopreservation is often ascertained by:

• Cell and tissue survival.
• The ability to re-grow or regenerate into complete plants or form new colonies [43].

It is desirable to assess the genetic integrity of recovered germplasm to determine whether it is ‘true-to-type’ following cryopreservation [44]. The fidelity of recovered plants can be assessed at phenotypic, histological, cytological, biochemical, and molecular levels, although there are advantages and limitations to the various approaches used to assess genetic stability [45]. Cryobionomics is a new approach to study genetic stability in cryopreserved plant materials [46]. The embryonic tissues can be cryopreserved for future use or for germplasm conservation [47].

Haploid Production for Plant Breeding

Tissue culture techniques enable the production of homozygous plants in a relatively short time period through protoplast, anther, and microspore cultures, offering an alternative to conventional breeding methods [53].

Haploids are sterile plants having a single set of chromosomes. These are converted into homozygous diploids by spontaneous or induced chromosome doubling. The doubling of chromosomes restores the fertility of plants, resulting in the production of double haploids with the potential to become pure breeding new cultivars [54].

The term androgenesis refers to the production of haploid plants from young pollen cells without undergoing fertilization. Sudherson et al. [55] reported haploid plant production of Sturt’s desert pea by using pollen grains as primary explants.

Haploidy technology has now become an integral part of plant breeding programs by:

1. Speeding up the production of inbred lines [56].
2. Overcoming the constraints of seed dormancy and embryo non-viability [57].

The technique has a remarkable use in genetic transformation by the production of haploid plants with induced resistance to various biotic and abiotic stresses. Introduction of genes with desired traits at the haploid state, followed by chromosome doubling, has led to the successful production of double haploids in inbred wheat and drought-tolerant plants [58].

Embryo Culture Techniques

Embryo culture is a type of plant tissue culture used to grow embryos from seeds and ovules in a nutrient medium. In embryo culture, the plant develops directly from the embryo or indirectly through the formation of callus and then subsequent formation of shoots and roots.

The technique has been developed to achieve several goals [59, 119]:

• Break seed dormancy.
• Test the vitality of seeds.
• Production of rare species and haploid plants.

It is an effective technique employed to shorten the breeding cycle of plants by growing excised embryos, resulting in the reduction of the long dormancy period of seeds. Intra-varietal hybrids of an economically important energy plant, Jatropha, have been produced successfully with the specific objective of mass multiplication [62].

Somatic embryogenesis and plant regeneration have been carried out in embryo cultures of Jucara Palm for rapid cloning and improvement of selected individuals [60]. In addition, conservation of endangered species can also be attained by practicing the embryo culture technique. Recently, a successful protocol has been developed for the in vitro propagation of Khaya grandifoliola by excising embryos from mature seeds [61]. This plant has a high economic value for timber wood and for medicinal purposes.

This technique has an important application in forestry by offering a means of propagation of elite individuals where the selection and improvement of natural populations is difficult.