Online Material
Supplementary Figures
Fig. S1. High resolution confocal images of anaphase and telophase stage nuclei from
Hrb87F-null embryo. (A) Side-by-side association of telomeres between chromatids extending
from the two poles of anaphase stage. (B-D) Telophase stage nuclei showing progressive
separation of the side-by-side associated chromatids. Nuclei are stained with DAPI. Arrows point
to the telomere associations.
Fig. S2. Nuclear divisions in Hrb87F-null early embryos are asynchronous. Unlike the
highly synchronous nuclear divisions in WT embryos (A), the Hrb87F-null early embryos show
considerable asynchrony in nuclear divisions along the antero-posterios axis (B): this example
shows late telophases at anterior and posterior ends (yellow line encircled), anaphases/early
telophases in the next two zones on either ends (red line encircled) and metaphases in the middle
zone (blue line encircled). Note the chromatin bridges (telomeric associations) between the two
poles in all anaphase and early telophase nuclei.
Supplementary Methods
Fly crosses to insert a normal telomere bearing X-chromosome (with or without the
hsFLAG-Hrb87F transgene) in tsr+/tsr+; +/+ or tsr+/tsr+; Df(3R)Hrb87F/Df(3R)Hrb87F) background
Scheme A. Generation of w hsFLAG-Hrb87F; tsr+; Df(3R)Hrb87F stock
Cross 1.
w hsFLAG-Hrb87F; +; + (♀)
X
Select red eye, Sp and non-Tb (♂) flies
w hsFLAG-Hrb87F; +/Sp; +/dco2 e
w1118; Sp/CyO; dco2 e/TM6B (♂)
Select red eye, Cy and Tb (♀) flies
w hsFLAG-Hrb87F/w1118; +/CyO; +/TM6B
Cross 2.
w hsFLAG-Hrb87F; +/Sp; +/dco2 e X
Select red eye, Sp, Cy, ebony and Tb (♂) flies
w hsFLAG-Hrb87F; Sp/CyO; dco2 e/TM6B
w hsFLAG-Hrb87F/w1118; +/CyO; +/TM6B
Select dark red eye, Sp, Cy, ebony and Tb (♀) flies
w hsFLAG-Hrb87F; Sp/CyO; dco2 e/TM6B (*)
Cross 3.
w hsFLAG-Hrb87F; Sp/CyO; dco2 e/TM6B (♂) X
w1118 #; tsr+; Df(3R)Hrb87F (♀)
(# indicates X with elongated telomere)
Select red eye, Cy and Tb (♂) flies
w1118 #; tsr+/CyO; Df(3R)Hrb87F/TM6B (**)
Cross 4.
w1118 #; tsr+/CyO; Df(3R)Hrb87F/TM6B(♂)(**) X
(♂ progeny from Cross 3 above **)
w hsFLAG-Hrb87F; Sp/CyO; dco2 e/TM6B (♀) (*)
(♀ progeny from Cross 2 above *)
Select red eye, non-Sp, Cy, non-ebony and Tb (♂) flies
w hsFLAG-Hrb87F; tsr+/CyO; Df(3R)Hrb87F/TM6B
Cross 5.
w hsFLAG-Hrb87F; tsr+/CyO; Df(3R)Hrb87F/TM6B (♂) X
w hsFLAG-Hrb87F; Sp/CyO; dco2 e/TM6B (♀) (*)
Select red eye, non-Sp, Cy, non-ebony and Tb ♂ and ♀ flies
w hsFLAG-Hrb87F; tsr+/CyO; Df(3R)Hrb87F/TM6B
Cross 6.
w hsFLAG-Hrb87F; tsr+/CyO; Df(3R)Hrb87F/TM6B X w hsFLAG-Hrb87F; tsr+/CyO; Df(3R)Hrb87F/TM6B
Select red eye, non-Cy, non-Tb flies
w hsFLAG-Hrb87F; tsr+/tsr+; Df(3R)Hrb87F/Df(3R)Hrb87F
(maintained as stock for examination of telomeres on different chromosomes in subsequent generations)
Scheme B. Generation of w1118; tsr+; Df(3R)Hrb87F stock
Cross 1.
w1118; Sp/CyO; dco2 e/TM6B (♀) X w1118 #; tsr+; Df(3R)Hrb87F (♂)
(# indicates X with elongated telomere)
Select red eye, Cy and Tb (♂) flies
1118
w
; tsr+/CyO; Df(3R)Hrb87F/TM6B
Cross 2.
w1118; tsr+/CyO; Df(3R)Hrb87F/TM6B (♂) X w1118; Sp/CyO; dco2 e/TM6B (♀)
Select red eye, non-Sp, Cy, non-ebony and Tb ♂ and ♀ flies
w1118; tsr+/CyO; Df(3R)Hrb87F/TM6B (***)
Cross 3.
w1118; tsr+/CyO; Df(3R)Hrb87F/TM6B X
w1118; tsr+/CyO; Df(3R)Hrb87F/TM6B
w1118; tsr+/tsr+; Df(3R)Hrb87F/Df(3R)Hrb87F
(maintained as stock for examination of telomeres on different chromosomes in subsequent generations)
Scheme C. Generation of w1118; tsr+; + stock
Cross 1.
w1118; Sp/CyO; +/+ (♀)
X w1118; tsr+/CyO; Df(3R)Hrb87F/TM6B (♂)
(male progeny from Cross 3 of Scheme B ***)
Select red eye, non-Sp, Cy and Tb flies
w1118; tsr+/CyO; +/TM6B (Self Cross)
Cross 2.
w1118; tsr+/CyO; +/TM6B (♀)
X w1118; tsr+/CyO; +/TM6B (♂)
Select red eye, non-Cy and non-Tb flies
w1118; tsr+/ tsr+; +/+
(maintained as stock for examination of telomeres on different chromosomes in subsequent generations)