Assignment

Plant cell and tissue culture

Topic:

Somaclonal variation as breeding tool

Somatic hybridization

Submitted to:

Dr. Sajjad

Submitted by:

Khadija Tahira

BBCH-01103011

Bs-Biochemistry

8th semester

Dated: 2nd April, 2014

Somaclonal variation:

Plants regenerated in vitro from undifferentiated cells often exhibit some level of variation called

somaclonal variation. Somaclonal variation in plants is caused by increased mutation rates

mainly due to nucleotide substitutions and small indels during the in vitro regeneration process

and could be provoked by exogenous growth stimulators. Thereby, genetic modifications lead to

somaclonal variations in regenerated plantlets (Volodymyr et al., 2012).

History:

It has been known for over 25 years now that plant cell and tissue cultures undergo genetic

erosions and show changes of various types, especially in chromosome numbers and ploidy level

(Partanen 1963; D’Amato 1965; Murashige and Nakano 1966). However, till the mid 1970’s

such changes were considered undesirable and were therefore discarded because the main

emphasis was on clonal propagation and genetic stability of the cell cultures. Extensive studies

conducted during the last decade have shown that the cell cultures, especially on periodical

subculturing, undergo various morphological and genetic changes, i.e., polyploidy, aneuploidy,

chromosome breakage, deletions, translocations, gene amplification, inversions, mutations, etc.

(see Nagl 1972; Meins 1983; D’Amato 1985(Y. P. S. Bajaj (1990)

Somaclonal selection utilizing tissue culture and directed mutagenesis is a rapid breeding

strategy that has the benefit of retaining many of the key attributes of the parental line in the

derived clones, unlike the genetic reassortment that occurs with traditional breeding sexual

crosses. This particularly has importance for the French fry processing sector which has highly

demands on varietal characteristics. Russet Burbank is over 130 years old and yet remains the

most commonly used variety for the French fry processing industry globally despite over a

century of breeding efforts. Somaclonal type selection methods have been used to improve

resistance to several important potato pathogens. In production of these variants the somaclonal

selection process targeted common scab resistance using the pathogen’s key pathogenicity

determinant, the toxin thaxtomin A, as a positive cell selection agent. The aim was to obtain

toxin-tolerant plants that express disease resistance. Strong and robust disease resistance was

obtained but not always in variants expressing toxin tolerance (Tamilarasan et al., 2013).

Somaclonal variation as a tool for crop improvement:

Somaclonal variation is a tool that can be used by plant breeders. The review examines where

this tool can be applied most effectively and the factors that limit or improve its chances of

success. The main factors that influence the variation generated from tissue culture are:

(1) The degree of departure from organized growth,

(2) The genotype,

(3) Growth regulators and

(4) Tissue source.

Despite an increasing understanding of how these factors work it is still not possible to predict

the outcome of a somaclonal breeding programme. New varieties have been produced by

somaclonal variation, but in a large number of cases improved variants have not been selected

because:

(1) The variation was all negative,

(2) Positive changes were also altered in negative ways,

(3) The changes were not novel, or

(4) The changes were not stable after selfing or crossing.

Somaclonal variation is cheaper than other methods of genetic manipulation. At the present time,

it is also more universally applicable and does not require ‘containment’ procedures. It has been

most successful in crops with limited genetic systems and/or narrow genetic bases, where it can

provide a rapid source of variability for crop improvement.(Angela karp 1995)

Somaclonal variation caused by in vitro culture is also called tissue culture-induced variation.

Somaclonal variation may involve chromosome number and structure, gene mutation, altered

sequence copy number, activation of transposable elements, somatic crossing-over, sister

chromatide exchange, DNA amplification and deletion, and change in methylation pattern.

Genotype and in vitro culture conditions (type of explants, medium, duration) influence the

occurrence and frequency of somaclonal variation (Paul et al., 2008).

Limitations to the Use of Somaclonal Variation in Crop Improvement:

Somaclonal variation in the major crop plants, rice, wheat, maize, barley, triticale, sugarcane,

potato and a few forage grasses is reviewed. Reported somaclonal variants include chlorophyll-

deficient plants, and those with changed morphology, single-gene mutations, polyploidy,

aneuploidy, chromosomal re-arrangements, modified yield, quality and disease resistance, and

occasionally novel variants not present in the natural gene pools. Somaclonal variation results

from both dominant and recessive mutations. The type and frequency of variants suggests that

somaclonal variation is akin to non-directed, random mutagenesis which generates a large

amount of unwanted variation. Consequently, most of somaclonal variation is either useless or of

limited use in direct varietal upgrading. However, somaclonal variants are easier to detect than

those in conventional mutagenesis. It is concluded that the development of in-vitro selection

procedures is essential to sieve out useful from useless variation to overcome the constraints of

somaclonal variation in breeding programs.(B. S. Ahloowalia 1986)

The origins of somaclonal variations are important for ensuring the validity of this research.

Generation of somaclonal variations is attributed to genetic and epigenetic modifications in

DNA. In particular, transposable elements (TEs) are one of the causes of genetic rearrangements

in invitro culture. Tissue culture is reported to activate silent TEs, resulting in somaclonal

variations (Mitsuru et al., 2011).

It is suggested that most somaclonal variations were derived from newly generated mutations

arising from the tissue culture process and not pre-existing mutations in the explants. Mutated

cell percentages remained low in the explants but increased rapidly during the tissue culture. In

recent years, molecular biological approaches have been utilized for detecting somaclonal

mutations, including amplified fragment length polymorphisms and intersimple sequence

repeats. These methods are preferable for detecting unidentified or unspecified mutations and

determining whether a mutation has occurred (Mitsuru et al., 2011).

Somatic hybridization:

Somatic hybridization in citrus: An effective tool to facilitate variety improvement:

Citrus somatic hybridization and hybridization via protoplast fusion has become an integral part

of citrus variety improvement programs worldwide. Citrus somatic hybrid plants have been

regenerated from more than 200 parental combinations, and several cybrid combinations have

also been produced. Applications of somatic hybridization to citrus scion improvement include

the production of quality tetraploid breeding parents that can be used in interploid crosses to

generate seedless triploids, and the direct production of triploids by haploid + diploid fusion.

Applications of somatic hybridization to citrus rootstock improvement include the production of

allotetraploid hybrids that combine complementary diploid rootstocks, and to combine citrus

with sexually.(J. W. Grosser et al 2000).

Since Carlson’s reports on the isolation of mutants and production of somatic hybrids from

tobacco tissue cultures, there has been interest in the development of somatic cell genetic

systems in plants. Increasing numbers of biochemically selectable markers have been reported in

the past ten years; protoplast fusion and plant regeneration have become useful tools for

generating interspecific and intergeneric hybrid plants. These techniques allow the combining of

genetic material from different somatic cells. In addition to permitting genetic exchange, an

efficient genetic system should provide opportunities for gene mapping, e.g., by somatic

recombination and chromosome elimination. Both somatic segregation and chromosome

instability have been observed in mammalian somatic hybrids. They also are associated with an

increase in ploidy level (Lazar et al .1981).

Plant regeneration and somatic embryogenesis through interspecific hybridization among

different Carica species were studied for the development of a papaya ringspot virus-resistant

variety. The maximum fruit sets were recorded from the cross of the native variety C. papaya cv.

Shahi with the wild species C. cauliflora. The highest hybrid embryos were recorded at 90 days

after pollination and the embryos were aborted at 150 days after pollination. The immature

hybrid embryos were used for plant regeneration and somatic embryogenesis (Latifah et al.,

2012).

There has been a lot of work to transfer potentially valuable disease-resistance genes from

diploid sexually incompatible Solanum species into the cultivated potato via somatic

hybridization. Potato protoplasts have been fused with a number of sexually incompatible wild

Solanum species (including S. hmidens, S. bulbocastanum, S. commersonii, S. polyadenium and

S. etuberosum), and many fertile somatic hybrid plants have been regenerated. Somatic hybrids

have been screened for disease resistances and these resistances are heritable (Mitchell et al.,

1995).

Refrences:

Voldodymyr, R., Radchuk, R., Pirko, Y., Vankova, R., Gaudinova, A., Korkhovoy, V., Yemets, A., Weber, H., Weschke, W., and Blume, Y. 2012. A somaclonal line SE7 of finger millet (Eleusine coracana) exhibits modified cytokinin homeostasis and increased grain yield. Journal of experimental botany 63:5497-5506.

Tamilarasan, T., Tegg, R., and Wilson, C. 2014. Resistance to Multiple Tuber Diseases Expressed in Somaclonal Variants of the Potato Cultivar Russet Burbank. The scientific world journal 2014: 8 pages.

Paul, S., Mohler, V., Wenzel, G., and Walter, B. 2008. Variation in DNA methylation patterns of grapevine somaclones (Vitis vinifera L.). BMC Plant biology

Mitsuru, S., Hosokawa, M., Doi, M. 2011. Somaclonal Variation Is Induced De Novo via the Tissue Culture Process: A Study Quantifying Mutated Cells in Saintpaulia. Plos one. n. page.

Chuanen, Z., Wei, D., Lu, H.,, Jiajie, W., Li, J., Yang, T., Daying, Z., Zeng-Yu, W., Guangmin, X. 2012. Construction of Whole Genome Radiation Hybrid Panels and Map of Chromosome 5A of Wheat Using Asymmetric Somatic Hybridization. Plos one. n. page.

Deval, P., J. Brian P., Paul, A., Farah, B., J. S. (Pat) Heslop-Harrison and Michael R. 2011. Somatic hybrid plants of Nicotiana 3 sanderae (1) N. debneyi with fungal resistance to Peronospora tabacina. Annals of Botany 108: 809–819.

Md. Abul, A., Md. Golam, R., and Latifah, A., 2012. Plant Regeneration and Somatic Embryogenesis from Immature Embryos Derived through Interspecific Hybridization among Different Carica Species. Int. J. Mol. Sci. 13:17065-17076.

J. Mitchell M, Susan and John P. 1995. Segregation and Recombination of Solanum bravidens Synteny Groups in Progeny of Somatic Hybrids With S. tuberosum: Intragenomic Equals or Exceeds Intergenomic Recombination. Genetics society of America 142: 1335-1348.

Y. P. S. Bajaj Biotechnology in Agriculture and Forestry Volume 11, 1990, pp 3-48

Angela karp Euphytica February 1995, Volume 85, Issue 1-3, pp 295-302

R. Bozorgipour, J.W. Snape Euphytica 04-1997, Volume 94, Issue 3, pp 335-340

B. S. Ahloowalia Advances in Agricultural Biotechnology Volume 20, 1986, pp 14-27

J. W. Grosser,  P. Ollitrault, O. Olivares-Fuster In Vitro Cellular & Developmental Biology - Plant November–December 2000, Volume 36, Issue 6, pp 434-449

Horst Binding, Reinhard Nehls Molecular and General Genetics MGG 17. VIII. 1978, Volume 164, Issue 2, pp 137-143

Jihong Liu, Xiaoyong Xu, Xiuxin Deng Plant Cell, Tissue and Organ Culture July 2005, Volume 82, Issue 1, pp 19-44

Deepak Pental, John D. Hamill, Andrew Pirrie, Edward C. Cocking Molecular and General Genetics MGG March 1986, Volume 202, Issue 3, pp 342-347