The below mentioned article provides the advantages of pollen culture over anther culture.
Anther culture is an efficient way for the production of haploids from the microspores present within the intact anther.
In this process, there is always the possibility that somatic cells of the anther that are diploid, will also respond to the culture condition and so produce unwanted diploid calluses or plantlets.
Sometimes, the development of microspore inside the anther may be interrupted due to growth inhibiting substances leaking out of the anther wall in contact with nutrient medium. In attempts to avoid these problems, the culture of free pollen has been investigated.
The culture of pollen offers the following additional advantages:
(i) Overcrowding of pollen grain in anther is eliminated and isolated pollen grains are equally exposed to nutrient medium.
(ii) Unwanted growth of the anther wall and other associated tissue are eliminated.
(iii) The steps of androgenizes can be observed starting from single cell.
(iv) Various factor governing androgenizes can be better regulated.
(v) Pollen is ideal for uptake, transformation and mutagenic studies as pollens can be uniformly exposed to chemicals and physical mutagens.
(vi) Pollen may be directly transformed into an embryoid. So it is very suitable for understanding biochemistry and physiology of androgenizes.
(vii) Higher yields of haploid plants per anther could be expected in pollen culture than the anther culture.
Haploid plants derived from either anther culture or pollen cultures are sterile. These plants contain only one set of chromosomes. By doubling their chromosome number, the plants can be made fertile and the resultant plants will be homozygous diploid or isogenic diploid (Fig 11.4). These homozygous diploid plants show the normal meiotic segregation. The fertile homozygous diploid plants are more important than the sterile haploid plants. Homozygous diploid plants can be used as pure lines in breeding programme.
Haploids can be diplodized by a number of methods:
Colchicine has been utilized widely as a spindle inhibitor to induce chromosome duplication and to produce polyploid plants. This method has been employed for obtaining homozygous diploid plants from haploid culture. The young, plantlets while still enclosed within anther, are treated with 0.5% colchicine solution for 24-48 hrs. Treated plantlets are replanted in the medium after thorough washing. In case of mature haploid plantlets, 4% colchicine-lanoline paste may be applied to the axil of the leaves.
It is known that haploid cells are unstable in culture and have a tendency to undergo endomitosis, i.e., chromosome duplication without nuclear division. This property can be used for obtaining homozygous diploid plants.
In this process, a small explant of stem from a haploid plant is cultured on auxin-cytokinin added medium where the segment forms the callus tissue. During callus growth, diploid homozygous cells are produced by endomitosis. Now large number of isogenic diploid plants can be obtained by organogenesis.
Fusion of Pollen Nuclei:
Homozygous diploid callus or embryoids may form by the spontaneous fusion of two similar nuclei of the cultured pollen after first division. In Brassica, the frequency of spontaneous nuclear fusion in microspore is high in culture.
Significance of Haploid Plants:
In a diploid cell the chromosomes exist in homologous pairs. The genes for specific characters are also formed in pairs which are known as allelic gene pairs. For an example, T gene (for tallness) is an allele of t gene (for dwarfness) and vice versa in heterozygous condition. Each allele is located on one of the pair of homologous chromosome at a particular gene locus.
Although each allele controls the same genetic trait (height of the plant), yet, they may control a contrasting phenotypic expression (tall/dwarf) of that trait. In heterozygous condition, the activity of only one of the alleles is expressed phenotypically, the allele is said to be dominant (suppose T gene). On the other hand, the activity of the other allele which is not expressed phenotypically until, it is separated from dominant allele, is said to be recessive (suppose t gene).
A chromosome contains a number of dominant and recessive genes whose allelic forms are present on the homologous partner chromosome at the same gene loci. In heterozygous diploid, only dominant alleles are expressed phenotypically. Homologous chromosome separates during meiosis. Pollen grains receive only one set of homologous chromosomes.
As a result, in pollen-derived haploid plants, all the recessive genes, along with dominant genes, will be expressed phenotypically as there is no masking of recessive gene by dominant genes. Since the haploid plants are sterile, diploid fertile plant can be made by doubling the same existing chromosome.
As a result, the dominant as well as recessive genes will be doubled at their respective loci. So, even in diploid condition, all the recessive genes will be expressed phenotypically. Such diploid plant is also called homozygous plant or isogenic diploid plant. Therefore, comparing the heterozygous diploid plant with homozygous diploid plant, one can easily identify the recessive characters which are not possible to identify in heterozygous condition.
Crossing over is an essential feature in the meiotic cycle by which random exchange of genetic material (genetic recombination) between two homologous chromatids takes place. The exchange is of great importance because it produces a new gene combination. As a result, four haploid nuclei, produced from a single diploid nucleus, differ from one another. Therefore, haploid plants, derived from four haploid pollens of a pollen tetrad, are significant because the plants will differ genotypically.
Why the Haploid Plants are Sterile?
In haploid plants, each chromosome is represented only once and this is the reason there is no zygotene pairing in first meiotic division. Thus, all the chromosomes appear as univalent. During anaphase I, each chromosome moves freely and form generally more than two groups. Gametes with less than the haploid number are generally not viable; therefore, haploid plants are highly sterile.