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<P>Tonami <I>et al</I>. 10.1073/pnas.0407236102.</TD>
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<H2>Supporting Information</H2>
<H4>Files in this Data Supplement:</H4>
<A HREF="#F1">Supporting Figure 6</A><BR>
<A HREF="#F2">Supporting Figure 7</A><BR>
<A HREF="#F3">Supporting Figure 8</A><BR>
<A HREF="#F4">Supporting Figure 9</A><BR>
<A HREF="#F5">Supporting Figure 10</A><BR>
<A HREF="#F6">Supporting Figure 11</A><BR>
<A HREF="#F7">Supporting Figure 12</A><BR>
<BR><BR><BR><BR>
<A href="{__picrender__}pnas_102_16_5797__1.pdf">Supporting Figure 6</A>
<A NAME="F1"></A>
<B><P>Fig. 6.</B> Ribonucleotide reductase (RNR) is required for double-strand break (DSB) formation. (<I>a</I>�<I>c</I>) DNA content of the cells in Fig. 1 <I>B</I>�<I>D</I> is shown in <I>a</I>�<I>c</I>, respectively. (<I>d</I>�<I>g</I>) <I>pat1 rec12</I> (HM336) (<I>d</I> and <I>e</I>) and <I>pat1 rec12 rad50S</I> (HM1805) (<I>f</I> and <I>g</I>) cells were arrested in G<SUB>1</SUB> phase, and meiosis was induced at 34�C in the absence (�) or presence (+) of 24 mM hydroxyurea (HU). DNA content of the cells in <I>d</I> and <I>f</I> is shown in <I>e</I> and <I>g</I>, respectively. (<I>d</I> and <I>f</I>) Samples were subjected to digestion of DNA with <I>Not</I>I and then analyzed by pulsed-field gel electrophoresis (PFGE) and Southern blot hybridization with the probe as in Fig. 1<I>A</I>.</P>
<BR><BR><BR><BR>
<A href="{__picrender__}pnas_102_16_5797__2.pdf">Supporting Figure 7</A>
<A NAME="F2"></A>
<B><P>Fig. 7.</B> RNR is required for DSB formation. (<I>a</I>) Position of the probe corresponding to the <I>rae1<SUP>+</I></SUP> fragment of <I>Not</I>I C fragment of chromosome 2. This probe was used in <I>b</I>�<I>e</I>. <I>pat1</I> (HM1307) (<I>b</I>), <I>pat1 rec12</I> (HM336) (<I>c</I>), <I>pat1 rad50S</I> (HM1691) (<I>d</I>), and <I>pat1 rec12 rad50S </I>(HM1805) (<I>e</I>) cells were arrested in G<SUB>1</SUB> phase, and meiosis was induced at 34�C in the absence (�) or presence (+) of 24 mM HU. Samples collected at the indicated times were analyzed as in Fig. 1<I>B</I>.</P>
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<A href="{__picrender__}pnas_102_16_5797__3.pdf">Supporting Figure 8</A>
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<B><P>Fig. 8.</B> Rad3p is required for coupling of meiotic DNA replication to DSB formation. (<I>a</I>�<I>h</I>) <I>pat1 rad3</I> (HM2877) (<I>a</I> and <I>b</I>), <I>pat1 rad3 rec12</I> (HM2849) (<I>c</I> and <I>d</I>), <I>pat1 rad3 rad50S</I> (HM2690) (<I>e</I> and <I>f</I>), or <I>pat1 rad3 rec12 rad50S</I> (HM2699) (<I>g</I> and <I>h</I>) cells were arrested in G<SUB>1</SUB> phase, and meiosis was induced at 34�C in the absence (�) or presence of HU (+), with HU added at time 0. Samples collected at the indicated times thereafter were analyzed as in Fig. 1<I>B</I>. The probe used in <I>a</I>, <I>c</I>, <I>e</I>, and <I>g</I> is the one shown in Fig. 7<I>a</I>. DNA content of the cells in <I>a</I> and Fig. 3<I>A</I>, <I>c</I> and Fig. 3<I>B</I>, <I>e</I> and Fig. 3<I>C</I>, and <I>g</I> and Fig. 3<I>D</I> is shown in <I>b</I>, <I>d</I>, <I>f</I>, and <I>h</I>, respectively.</P>
<BR><BR><BR><BR>
<A href="{__picrender__}pnas_102_16_5797__4.pdf">Supporting Figure 9</A>
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<B><P>Fig. 9.</B> Cds1p is required for coupling of meiotic DNA replication to DSB formation. (<I>a</I>�<I>e</I>) <I>pat1 cds1 rad50S</I> (HM1801) (<I>a</I> and <I>b</I>) and <I>pat1 cds1 rec12 rad50S</I> (HM1842) (<I>c</I>�<I>e</I>) cells were arrested in G<SUB>1</SUB> and induced to undergo meiosis at 34�C, and samples were analyzed as in Fig. 1<I>B</I>. The probe used in <I>a</I> and <I>d</I> was the one shown in Fig. 7<I>a</I>, whereas that in <I>c</I> was in Fig. 1<I>A</I>. DNA content of the cells in <I>a</I> and Fig. 4<I>B</I> and <I>c</I> and <I>d</I> is shown in <I>b</I> and <I>e</I>, respectively.</P>
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<A href="{__picrender__}pnas_102_16_5797__5.pdf">Supporting Figure 10</A>
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<B><P>Fig. 10.</B> Cdc2p/Cdc13p is not involved in coupling meiotic DNA replication to DSB formation. (<I>a</I> and <I>b</I>) DNA content of the cells in Fig. 5 <I>A</I> and <I>B</I> was shown in <I>a</I> and <I>b</I>, respectively. (<I>c</I>�<I>f</I>) <I>pat1 rad3 rec12 cdc2-22</I> (HM4082) (<I>c</I>) and <I>pat1 rec12 cdc2-22</I> (HM4133) (<I>d</I>) cell were arrested in G<SUB>1</SUB> phase, and meiosis was induced at 36.5�C in the absence (�) or presence (+) of HU, with HU added at time 0. Samples collected at the indicated times thereafter were analyzed by PFGE as in Fig. 1<I>B</I>. DNA content of the cells in <I>c</I> and <I>d</I> is shown in <I>e</I> and <I>f</I>, respectively. The probe used <I>c</I> and <I>d</I> was the one shown in Fig. 1<I>A</I>. (<I>g</I> and <I>h</I>) DNA content of the cells in Fig. 5 <I>C</I> and <I>D</I> is shown in <I>g</I> and <I>h</I>, respectively. (<I>i</I>�<I>l</I>) <I>pat1 rad3 rec12 cdc13 rad50S</I> (HM4130) (<I>i</I>), <I>pat1 rec12 cdc13 rad50S</I> (HM4132) (<I>k</I>) cells were arrested in G<SUB>1</SUB> phase, and meiosis was induced at 34�C in the absence (�) or presence (+) of HU, with HU added at time 0. Thiamine was added at time 0 to switch off <I>cdc13<SUP>+</I></SUP> expression. (<I>i</I> and <I>j</I>) Samples collected at the indicated times thereafter were analyzed by PFGE as in Fig. 1<I>B</I>. (<I>k</I> and <I>l</I>) DNA content of the cells in <I>i</I> and <I>j</I> is shown in <I>k</I> and <I>l</I>, respectively. The probe used in <I>i</I> and <I>j</I> is the one shown in Fig. 1<I>A</I>.</P>
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<A href="{__picrender__}pnas_102_16_5797__6.pdf">Supporting Figure 11</A>
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<B><P>Fig. 11.</B> Tyrosine phosphorylation of Cdc2p is not required for coupling meiotic DNA replication to DSB formation. (<I>a</I>�<I>h</I>) <I>pat1 mik1 wee1</I> (HM6 05) (<I>a</I>) <I>pat1 mik1 wee1 rad50S</I> (HM1804) (<I>c</I> and <I>e</I>), and <I>pat1 mik1 wee1 rec12 rad50S</I> (HM1824)(<I>f</I> and <I>h</I>) cells were arrested in G<SUB>1</SUB> phase, and meiosis was induced at 34�C in the absence (�) or presence (+) of HU, with HU added at time 0. Samples collected at the indicated times thereafter were analyzed by PFGE as in Fig. 4<I>A</I> for <I>a</I> and as in Fig. 1<I>B</I> for <I>c</I>, <I>e</I>, <I>f</I>, and <I>h</I>. The probe used in <I>c</I> and <I>f</I> is the one shown in Fig. 1<I>A</I>, whereas that in <I>e</I> and <I>h</I> is in Fig. 7<I>a</I>. DNA content of the cells in <I>a</I>, <I>c</I> and <I>e</I>, and <I>f</I> and <I>h</I> is shown in <I>b</I>, <I>d</I>, and <I>g</I>, respectively.</P>
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<A href="{__picrender__}pnas_102_16_5797__7.pdf">Supporting Figure 12</A>
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<B><P>Fig. 12.</B> Relation among Orp1p, Cdc2p and HU treatment. (<I>a</I>�<I>d</I>), <I>pat1 orp1</I>(HM338) (<I>a</I>) and <I>pat1 cdc2-L7</I> (HM113) (<I>b</I>) cells were arrested in G<SUB>1</SUB> phase, and meiosis was induced at 37�C in the absence (�) or presence (+) of HU, with HU added at time 0. Samples collected at the indicated times thereafter were analyzed by PFGE as in Fig. 4<I>A</I>. Ch1, Ch2 and Ch3 indicated the positions of chromosomes 1 (5.7 Mb), 2 (4.6 Mb), and 3 (3.5 Mb), respectively. The smear (*) shows DSBs generated during meiosis. DNA content of the cells in <I>a</I> and <I>b</I> is shown in <I>c</I> and <I>d</I>, respectively.</P>