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Pollination and fertilization under in vitro condition offer the opportunity for producing hybrid embryo that not possible by conventional method . INTRODUCTION

What is mean of in vitro pollination and fertilization?:

Def:- ‘‘Under controlled condition aseptic transfer of pollen grain on stigma to produce hybrid embryos among plants that can’t cross by conventional method of plant breeding is called as in vitro pollination and fertilization ’’ What is mean of in vitro pollination and fertilization?


In nature intergenic and intraspecific hybridization occurs less frequently , this is due to hindering factor . Factor:- Barrier to growth of pollen tube on the stigma or style Solution:- In such cases the style or part of it can be excised and pollen grain either placed on the cut surface of the ovary IMPORTANCE


This technique is called as intraovarian pollination . Ex. Papaver somnifera, Argemone mexicana, etc.. To overcome the barriers to pollen tube growth is direct pollination of culture ovules Continue…


1.Stigmatic pollination :- application of pollen to the stigma in vitro condition . 2.Ovarian pollination :- application of pollen to exercised ovary in vitro condition . 3.Placental pollination :- application of pollen to ovules attached to the placenta in vitro condition. 4.Ovular pollination :- application of pollen to excised ovules in vitro condition. DIFFERENT TECHNIQUES


The ovaries which are large and which contain many ovules . Other essential materials is pollen which should be viable and able to germinate . 1% calciumchloride solution that favour the growth of pollen tube. MATERIAL REQUIRED


Flower bud are emasculated before anthesis and bagged in order to prevent the undesirable pollination the whole pistil or ovaries alone are sterilized by a quick rinse in 70% alcohol, surface sterilized with suitable agents. To collect the pollen under aseptic condition anther removed from bud or open flower. then kept in sterile Petri dish contain presterilised filter paper until their dehiscence.The dehiscence pollen is asceptically deposited on the cultred ovules, placenta or stigma depend on nature of experiment. DISINFECTION OF MATERIAL

Culture of ovules and ovary:

Ovules:- The growth of pollen tubes attached to bare ovules is often inhibited by the presence of water on the surface of ovules this water is removed by filter paper and later dried ovules covered by pollen grain ,raising seed by ovules which contain globular embryo after in vitro pollination ovules contains a single celled zygote which requires more complex growth condition . For developing subsequently embryonic stages ovules which have been self pollinated are kept on placenta until seed form. Ex.Nicotina tabacum , Alliums cepa,etc.. Ovary:-this technique developed by Nitsch in 1951 who successfully reared ovaries excised from pollinated flower in vitro to develop into mature fruits. Ex. Linaria macroccana, Tropaeohim majus ,etc.. Culture of ovules and ovary


Intergenic hybridization . Intraspecific hybridization . Interspecific hybridization. Intrafamilar crossing. PURPOSE

Embryo rescue to overcome incompatible interspecific hybridization in Gossypium spp. Abstract :-:

Successfully introgression of desirable character from certain spp. Could not be achieved because of different incompatibility. Such incompatibility can be overcome by ovuloembryo culture. From this literature it appear that the technique involve in excision of 10 to 15 days old embryo of Gossypium spp. Which is culture in BT and MS media supplements with phytohormones. E mbryo rescue to overcome incompatible interspecific hybridization in Gossypium spp. Abstract :-


Cotton is leading fibre crop of the worlds belonging to the genus Gossypium which consist of 49 identified and described species, of these 44 spp are Diploid and remaining are allotetraploids. Certain crosses and the many combination are reported to be incompatible due to the failure of fertilization between the species of Gossypium. Several worker have been critically reviewed the compatibility relationship between various Gossypium spp. Introduction

Techniques of Embryo culture:

For excision of embryo and their transfer to nutrien medium asceptic procedure are necessary since the embryo are lodged in sterile environment of ovule Ovule are generally surface sterlized before soaking Then they are split open and embryo are transfer to the nutrient media The most important aspect of embryo culture is the selection of medium necessary to sustain its contiue growth Mature embryo are grown in inorganic salt medium suppement with sucrose Techniques of Embryo culture


Different vitamin, amino acid and hormone are required. Then plantlets are remove from original medium and nurtured in sterlized soil. Contd….

Invtro fertilization and ovule culture:

Its followed by culturing fertilized ovule to maturity is an important techniques employed to overcome the barrier inhibiting pollentube growth and very early stage embryo abortion. It is the promising approach and viable altrnative to somatic cell hybridization. Invtro fertilization and ovule culture

Factor affecting embryo culture:

Temperature – At higher temperature abscission of ball occurs within 4-5 days after pollination. Basal media- Growth of ovules is better in BT media as compared to White and MS media. Genotype – Embryo culture response depend upon genotype of parent. Factor affecting embryo culture

Conclusion :

Ovule culture is important tools for overcoming incampatibility amongs parents and the succesful obtaining hybrids from difficult distant cross combnitation Thus, from wild Gossypium spp to cultigen, interogression of desired genes is made possible. Conclusion


Overcoming self incompatibility. Overcoming the cross incompatibility. Haploid production through parthenogenesis. APPICATION OF IN VITRO POLLINATION AND FERTILIZATION


Journal Indian biotechnology, January 2004 References:-

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