Chromosome doubling in haploids through colchicine
--P. Gayen, Jasbir Kaur Madan, Rajesh Kumar and K. R. Sarkar

Maternal haploids isolated through the genetic selection technique were used for chromosome doubling. Seeds (soaked for 48 hours) and seedlings (5 days after germination) were treated with 0.03%, 0.06% and 0.1% aqueous solution of colchicine (SRL) with DMSO (dimethyl sulfoxide) 0.5% (by volume) for 6 hours, 12 hours and 24 hours.

Seed treatment. A small portion of the plumule tip was cut off before treatment to ensure better penetrance of colchicine to the growing meristem. A set of controls without cutting the plumule tip was also included. Treatment with 0.06% colchicine concentration was carried out at two different temperatures, 18±1 C and 25±2 C. Fifty seeds per treatment were dipped in aqueous solutions of colchicine and kept in the dark. After completion of the treatment, the seeds were thoroughly washed with distilled water and sown in the field. Light watering in the evening up to 7 days after sowing is very essential for better germination and establishment of seedlings.

Seedling treatment. Shoot tips were nipped off to expose the apical meristem for better colchicine action. A set of controls without cutting the shoot tip was also included. Fifty seedlings per treatment were maintained by moistening cotton wads at 1 hour intervals. After completion of the treatment the shoot tips were thoroughly washed with distilled water. The seedlings were maintained in paper cups for 2-3 days in shade and then transferred to the field. Significant success (18.05%) was obtained from seed treatment (with plumule tip cut) with 0.06% colchicine concentration at 18±1 C for 12 hours (Table 1). Cutting of the plumule tip in the case of seed, and shoot tip in the case of seedlings, gave better response. No plant establishment was obtained from treatments with 0.06% and 0.1% concentrations for 24 hours. Chromosome doubling of treated haploids was confirmed by chromosome counting at diakinesis and anaphase I from the fixed male inflorescence.

Table 1. Effect of varying colchicine concentrations on doubling of maize haploids obtained through 'Scutellum Marker Technique.'
 
% doubled haploids
Duration of treatment
Colchicine Conc.
Material
6 hrs
12 hrs
24 hrs
0.03% Seedling Tip cut
1.02
1.20
1.80
Tip intact 0.00 0.29 0.50
Seed Tip cut 0.60 0.42 0.98
Tip intact 0.00 0.00 0.21
0.06% Seedling Tip cut 6.23 8.24 No plant estab.
Tip intact 3.29 4.26 -do-
Seed Tip cut 18 C 10.23 18.05 -do-
Tip cut 25 C 5.98 12.21 -do-
Tip intact 2.97 6.22 -do-
0.1% Seedling Tip cut 6.42 -do-
Tip intact 3.41 -do-
Seed Tip cut 4.32 5.38 -do-
Tip intact 2.21 3.61 -do-


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