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In Vitro Regeneration from Mature Embryos in Spring Wheat

M. M. Rahman
Scientific Officer
(Plant Breeding), Pulses Research Centre, Regional Agricultural Research Station, BARI, Ishurdi, Pabna

A. K. M. Shamsuddin
Professor
Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh

U. Asad
Scientific Officer
(Plant Breeding), Oilseed Research Centre, Regional Agricultural Research Station, BARI, Ishurdi, Pabna

Abstract/ Executive Summary:

The experiment was carried in laboratory of the Department of Genetics and Plant Breeding, BAU, Mymensingh during 2006. In vitro regeneration in wheat plants was carried out from mature embryos in five genotypes of spring wheat including two doubled haploids. Three concentrations of 2, 4-D were used for induction of callus. Maximum number of callus were produced on MS medium supplemented with 6.0 mg/L of 2, 4-D. But comparatively larger calli were produced on MS medium supplemented with 4.0 mg/L of 2, 4-D. Only two doubled haploid (DH-2 and DH-10) responded to regeneration. The genotype DH-10 performed better for callusing. Maximum numbers of plants were regenerated on the MS medium supplemented with 1.0 mg/L of kinetin which was followed by 2.0 mg/L of kinetin in the medium. Both the lines had similar regeneration capacity. Early shoot regeneration was observed on MS medium supplemented with 200.0 mg/L casein hydrolyses.

Keywords: Doubled haploid, Mature embryo, Regeneration, in vitro, Wheat

Location: Tthe green house and tissue culture laboratory, Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh

Start Date: 00-00-2006

End Date: 00-00-2006

Program Area: Variety and Species

Commodity: Wheat

Objectives:

To develop a protocol for regeneration of plants from mature embryos was attempted in five genotypes of wheat including two doubled haploids.

Methodology:

The experiment was carried in the green house and tissue culture laboratory of the Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh during 2006. Mature wheat (Triticum aestivum L.) embryos of five genotypes including two doubled haploid (DH) lines were used as explants for callus induction and subsequent plant regeneration. The genotypes were Sourav, Gourab, Shatabdi, DH-2 and DH-10. Mature seeds were soaked in petridish on a moistened filter paper for 2 hours before embryo dissection, so that the embryos can swell and become distinctly visible. The soaked seeds were sterilized by treating them with 70% ethanol for 15 minutes followed by washing with distilled water. This was followed by treating for 5 min in 30% chlorox (containing 4.7% chlorine). During sterilization the seeds were occasionally agitated. Finally the seeds were washed 5 times with sterile distilled water for removing the sterilant. Then the mature embryos were aseptically excised from the caryopsis and placed on callus induction medium in sterile petridish keeping the scutellum side up. Ten mature embryos were placed in each petridish and were sealed with parafilm . The callus induction media contained agar solidified MS media supplemented with three different doses of 2,4-D (4.0 mg/L, 5.0 mg/L, 6.0 mg/L, respectively). The explants were incubated in dark under 25 ±10C for 3-5 days. When callus initiation began the petridish were transferred to light with 16 hours photoperiod under the same temperature regime. The cultures were cheeked daily to note the initiation and the development of calli. After 30 days of inoculation, the developed calli were transferred to regeneration media. The regeneration media contained agar solidified MS media supplemented with different combinations and concentrations of growth regulators (MS as control, MS+ 1.0 mg/L of BAP, MS+ 2.0 mg/L of BAP, MS + 1.0 mg/L of kinetin, MS+ 2.0 mg/L of kinetin, MS + 100.0 mg/L of Casein hydrolysate and MS + 200.0 mg/L of Casein hydrolysate). All the explants, calli cultures were kept at 25±1°C under 3000 Lux light intensity and 16 hours photoperiod. The data were recorded for days to callus induction, number of callus initiated, callus size (mm), days to plant regeneration and number of plants regenerated. Data were estimated as percentage and the mean values in percent were used for analysis of data. The percent of callus induction and regeneration were estimated on the basis of the number of embryo and calli, respectively.

Source of Information: Int. J. Sustain. Crop Prod. 3(2):76-80, 2008

Article Link (Online Journal):

Funding Source:

1. Government

Budget:

Total Budget (Tk.) :

Achievement/Findings:

It could be concluded that good and rapid shoot regeneration was observed on medium containing 200.0 mg/L casein hydrolyses. On the other hand higher number of plant regeneration was observed on media supplement with kinetin although it took longer time.

Knowledge Management: Journal