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Modified Acetocarmine/n-banding Technique for Wheat Chromosome Identification at Meiosis

M. wazuddin
Dept. of Gentles and Plant Breeding, Bangladesn Aqricuhural Unlversltv, Mymenslngh.

Abstract/ Executive Summary:

A modified Acetocerrnlne/N - Banding technique have been developed and used to identify wheat chromosomes at meiosis. At meiotic metapase 1, nine of the 21 bivalents in Cornerstone, a hexaploid wheat have been recognized. The modified method produced more reliable stainjng and better chromosome definition.

Keywords: N-banding, Chromosome identification, Meiosis

Location: BAU, Mymensingh

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Wheat

Objectives:

To identify all nine pairs of identifiable chromosomes of hexaploid wheat in meiotic preparations.

Methodology:

The metaphase-1 chromosomes of pollen mother cells were investigated in Cornerstone- diploid wheat addition lines namely, m4. u4. th4 and u4 a: - Iso monosomic additlons. These addition lines are expected to have 21 pairs of cornerstone and either an entire, an iso or a telocentric form of the critical diploid chromosome. For modified ecatocarmine/N-bandlnq analysis one anther from each floret was used to check the meiotic metaphase-1 stage. The remaining two anthers were fixed In 3:1 ethanol : acetic acid fixing solution and stored at -15⁰C until used. The anther was macerated in 1 % aceto- carmine solution. heated gently and squashed. After removal of the cover- slip by liquid nitrogen, the slides were treated in 45% acetic acid at 55-60°C for 10-15 min. The slides were then air dried overnight. Next. the slides were incubated in hot phosphate buffer, 1M NaH₂P0₄at 90± 2^oC for'.5-2.0 min; and rinsed briefly in tap water. Slides were then stained in buffered 1 % (v/V) giemsa (Sigma) solution (10-20 min.' The slides were rinsed briefly in water. air dried. and stored. For recording N-bands. a coversiip was mounted on a crop of synthetic mounting medium, and the slides were observed under immersion oil. The N-banding method used in this programme is a modification of the technique developed by Endo and Gill (1984). The modifications were: (1) shorter. incubation in hot phosphate bufier. (ii) shorter staining time (10-12 rnin.), (iii) Squahing in 0.5 or 1% acetocarmine instead of longer pretreatment with 1% acetocarmine. Also. the giemsa stain (Sigma) was different from others. Giemsa stain used in this study was prepared by dissolving 10gm of giemsa stain' (Sigma) in 66 ml methanol + 66ml glycerol. The solution was stirred for about two hours at 60..± 2°C and then kept In dark at room temperature.

Source of Information: Bangladesh J. Agril. Sci. 17 (2) ; 277-280 : July 1990 : ISSN 0379-4296

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

At meiotic metapehase-1. nine of the 21 bivalents in 8 hexaploid wheat Cor- nerstone line exhibited distinct banding patterns. Rest of the Cornerstone and diploid chromosome 4(d4) added to Cornerstone did not show any recognizable bands. The modified method produced more reliable stainjng and better chromosome definition.

Knowledge Management: Journal