Nature - USA (2020-06-25)

Extended Data Fig. 7 | EdU pulse–chase analysis of endosymbiotic cells.
a, Pulse–chase experiments. The regeneration stalk was labelled with EdU at
regeneration day 3 and day 4. After washing out EdU, the samples were
cultured, collected and analysed at the indicated days during chasing.
b, Dissociated cells were processed by Click-iT to visualize EdU and stained with
DAPI to label nuclei. Cells were sorting on the basis of DAPI. c, DAPI-positive
cells were further sorted on the basis of Cy5.5 to estimate the number of the
total alga-free Xenia cells. d, e, To estimate the number of alga-containing Xenia
cells, free algae and alga-containing Xenia cells (alga+ population) were first
separated from all the other Xenia cells on the basis of the Cy5.5 signal (d). The
alga+ population was further separated into alga-containing Xenia cells and
free algae based on the SSC and FSC signals (e). f, g, Microscopy confirmation
of free algae (f) and alga-containing Xenia cells (g) sorted in e. Scale bars,
20 μm. In c–h, four independent experiments were carried out. h, The number
of EdU-positive and alga-containing Xenia cells were further estimated on the
basis of their strong EdU signal. i, Box plot of the percentage of EdU-positive
and alga-containing Xenia cells among all alga-containing Xenia cells at the
indicated days of chase. j, Box plot of the percentage of all alga-containing
Xenia cells among all Xenia cells at the indicated days of chase. Each dot in i, j
stands for one regenerating sample. Day 7, n = 3 polyps; day 9, n = 8 poly ps;
day 11, n = 6 polyps; day 13, n = 5 polyps; day 15, n = 3 polyps; day 17, n = 3 poly ps;
day 19, n = 3 polyps from 2 independent experiments were assayed. The
medians are indicated as lines in the boxes; the upper and lower edges of the
boxes represent the upper and lower quartiles, respectively.