Amplification of moderately repetitive DNA sequences during chick cartilage differentiation

Abstract

A 5-bromo-2'-[3H]deoxyuridine (BrdUrd) probe was isolated to analyze DNAs obtained from various chick tissues and cell types. [3H]BrdUrd-substituted DNA, prepared from limb bud cultures, was sheared and freed from palindromic DNA. Nonradioactive DNA was prepared from embryonic liver, undifferentiated limb bud mesenchyme, sternal cartilage, differentiated limb bud cultures, and BrdUrd-blocked cultures, and was sheared. These DNAs were used in 100-fold excess to drive the reassociation of the [3H]-BrdUrd-DNA probe. Purified mature cartilage DNAs of embryonic sternae or differentiated limb bud cultures drove the reassociation of the probe approximately two times faster than did DNA from liver, undifferentiated limb bud, or BrdUrd-blocked cells. These data indicate that cartilage DNA contains a greater number of sequences complementary to the BrdUrd probe than do DNAs of noncartilage or undifferentiated precartilage cells. Calculations determined an average substitution of 10% of thymidine residues by BrdUrd in purified probe, whereas CsCl density gradients of unsheared probe revealed radioactive peaks of greater than 20% substitution. The BrdUrd appears to be clustered in the genome.