Fig. 4.

Inhibition of hairpinning by RAG1 and RAG2 C-terminal regions. (A) Equal amounts of core RAG1/2 and noncore combinations were included in coupled cleavage reactions (12RSS/23RSS and Mg²⁺). The N and HP cleavage products from coupled cleavage reactions were analyzed by TBE-urea gel. (Right) Bar chart displays HP, N (percentage of total substrate + SD) and HP conversion factors (HP formed/total N produced + SD) from 3 independent experiments. (B) TBE-urea gel of hairpinning by various forms of RAG1/2 proteins activity on prenicked substrates, under coupled cleavage conditions. The arrowhead marks the position of shorter, inaccurate cleavage products resulting from alternative hairpinning sites opposite to the nick. Bar chart display the yield of double-strand breaks (DSB) produced from the total substrate as a percentage. The average was obtained from 3 experiments (+SD). (C) TBE-urea gel of hairpinning of prenicked substrates using the less stringent Mn²⁺ catalytic ion. Average percent DSB formed from total substrate are displayed in the bar chart (+SD).