I would like to add information about the technique of chromosome preparation in vivo, though this technique is not applicable to tissue culture or in vitro. However, researchers can follow this technique to standardise the chromosome preparation in vitro conditions. A good, condensed, deeply stained, and well-spread chromosome is essential to constructing a karyotype.

Chromosome preparation from bone marrow cells of mice or rats or even in tumour-bearing mice in vivo was performed by following the mitotic division inhibition technique (Chakrabarti et al., 1985; Mallick et al, 2018; Banerjee et al, 2019; Chowdhury and Banerjee, 2020;21). The technique is brief: i) Specimens received an injection of 0.04% colchicine solution intraperitoneally at a rate of 1 ml/100 g body weight for 1 hour 30 min. ii) Cells from bone marrow were collected in hypotonic solution (0.075MKCl) and aspirated. Then, the cell suspension was incubated for 30 min at 37 °C and centrifuged for 10 min at 1500 rpm to collect the cell sediment. The sediment was fixed in aceto-alcohol fixative (3:1, methanol: glacial acetic acid, v/v) and centrifuged for 8 min. The sediment was fixed again and kept for chromosomal slide preparation. The flame-dried chromosome-prepared slide was stained with 5% Giemsa diluted in phosphate (pH - 7) buffer for 50 min. Then slides were washed in running tap water. Well-spread metaphases were observed and analysed under the Binocular Research Microscope (10 100 magnifications) to construct karyotypes for studying chromosomal aberrations.