Feulgen microspectrophotometry, for the quantitative determination of DNA in situ, has been widely employed in plant (Kubica 1981, Price and Bachman 1975), animal (Pedersen et al. 1975) and human tissues (Lauder et al. 1978). Several positive correlations have been recorded between the nuclear DNA content and other nuclear characters (Evans et al. 1972, Jones and Rees 1968, Kulfinski 1975, Taper and Grant 1973). A direct proportionality between cytophotometric determination of DNA and histone content has also been observed (Sarma 1982) and additional genetic materials in chromosomes have been discussed in detail by Sharma (1983, 1984). The evolutionary significance of nuclear DNA content is now being widely discussed (Price 1976, Price and Bachmann 1975, Rees and Narayan 1977). It is suggested that both increase and decrease in DNA content may be associated with evolution. Haapala (1984) made an attempt to relate chromomeric loops and their interloop spaces to understand DNA sequence organization. In view of the importance of DNA content in evolution and its correlations with other nuclear characters as suggested for nucleotypic DNA, an attempt has been made for a critical analysis of intervarietal and interspecific variation, in Hordeum, in 4C nuclear DNA with the aid of Feulgen microspectrometry using two wavelength method. © 1987, Japan Mendel Society, International Society of Cytology. All rights reserved.