dark bands are darkly stained and the inter bands are lightly stained with nuclear stains. The dark bands contain more DNA and less RNA. The interbands contain more RNA and less DNA. The amount of DNA in interbands ranges from 0.8 - 25%. The bands of polytene chromosomes become enlarged at certain times to form swellings called puffs. The formation of puffs is called puffing. In the regions of puffs, the chromonemata uncoil and open out to form many loops. The puffing is caused by the uncoiling of individual chromomeres in a band. The puffs indicate the site of active genes where mRNA synthesis takes place. The chromonemata of puffs give out a series of many loops laterally. As these loops appear as rings, they are called Balbiani rings after the name of the researcher who discovered them. They are formed of DNA, RNA and a few proteins. As they are the site of transcription, transcription mechanisms such as RNA polymerase and ribonucleoproteins are present. In protozoans, there is no transcription, since the puff consists only of DNA. History Polytene chromosomes were originally observed in the larval salivary glands of Chironomus midges by Édouard-Gérard Balbiani in 1881. Balbiani described the chromosomal puffs among the tangled thread inside the nucleus, and named it "permanent spireme". In 1890, he observed similar spireme in a ciliated protozoan Loxophyllum meleagris. The existence of such spireme in Drosophila melanogaster was reported by Bulgarian geneticist Dontcho Kostoff in 1930. Kostoff predicted that the discs (bands) which he observed were "the actual packets in which inherited characters are passed from generation to generation." The hereditary nature of these structures was not confirmed until they were studied in Drosophila melanogaster in the early 1930s by German biologists Emil Heitz and Hans Bauer. In 1930, Heitz studied different species of Drosophila (D. melanogaster, D. simulans, D. hydei, and D. virilis) and found that all their interphase chromatins in certain cells were swollen and messy. In 1932, he collaborated with Karl Heinrich Bauer with whom he discovered that the tangled chromosomes having distinct bands are unique to the cells of the salivary glands, midgut, Malphigian tubules, and brain of the flies Bibio hurtulunus and Drosophila funebris. The two papers were published in the early 1933. Unaware of these papers, an American geneticist Theophilus Shickel Painter reported in December 1933 the existence of giant chromosome in D. melanogaster (followed by a