DEV021949 Supplementary Material

Files in this Data Supplement:

• Supplemental Table S1 -

  Adobe PDF

• Supplemental Figure S1 -

  Fig. S1. RNA in situ hybridisation patterns for all of the comets and cups. RNA in situ hybridisation to wild-type testes of all of the comet (upper panels) and cup (lower panels) genes. All testes are oriented with their apical tips towards the right-hand side.

• Supplemental Figure S2 -

  Fig. S2. Q-RT-PCR reveals that the comets and cups are transcribed before transition protein association with chromatin. (A) Q-RT-PCR profiles for control genes (CG3927, CG11591 and CG15177) for cysts from Tpl94D-EGFP, H2A-mRFP1 flies. (B) Q-RT-PCR profiles for comets and cups (hale, schuy, sunz, p-cup, d-cup, wa-cup and r-cup) for cysts from Tpl94D-EGFP, H2A-mRFP1 flies. In all cases the peak of post-meiotic transcription was detected in cysts positive for nuclear H2A-mRFP1, but lacking Tpl94D-EGFP. (C) Phase contrast, and H2A-mRFP (histone) and Tpl94D-EGFP (transition protein) fluorescent images of the cysts used for the Q-RT-PCR analysis. Asterisks indicate the nuclear ends of the elongated bundles.

• Supplemental Figure S3 -

  Fig. S3. Rfx is not the comet and cup transcription factor. Comet (hale, schuy, cola and soti) and cup (c-cup, w-cup, p-cup and wa-cup) transcript localisation is unaltered in Rfx⁴⁹/Rfx²⁵³ testes when compared with in wild type.

• Supplemental Figure S4 -

  Fig. S4. Hsr-omega transcripts are not detected in spermatids by in situ hybridisation. RNA in situ hybridisation to testes revealed that Hsr-omega is expressed in spermatogonia and young primary spermatocytes. The signal declined as primary spermatocytes matured, and the transcript was not detected in post-meiotic cells. Heat-shock treatment increased the signal intensity but the pattern remained unaltered. mRNA for the control gene CG33340 (hybridisation performed in parallel) was detected in primary spermatocytes and persisted into spermatid stages.

• Supplemental Figure S5 -

  Fig. S5. Transcript localisation and post-meiotic transcription are correlated in the CG11635-CG8701 cluster. (A) Genomic structure of the cluster, consisting of ten related genes arranged over 15 kb on chromosome 2R. cola and swif are annotated as a dicistronic transcript in FlyBase; however, we could not detect this dicistronic transcript by RT-PCR, and have annotated each as a separate gene. (B) In situ hybridisation patterns for all of the genes in the cluster. Four are expressed in primary spermatocytes, and the transcripts are stabilised through elongation (not comets, pale blue), while the other six show a comet transcript distribution (dark blue). (C) Q-RT-PCR results show that post-meiotic transcription correlates with the comet transcript localisation pattern; swif, boly and cola closely resemble hale, rather than sunz, in profile, while CG2127 was detected in all isolated cysts.

• Supplemental Figure S6 -

  Fig. S6. Q-RT-PCR data for sowi and soti. (A) Q-RT-PCR confirms that sowi resembles the sunz-group genes c-cup and d-cup. (B) We confirmed that soti is transcribed post-meiotically in a separate experiment.