def __init__(self, cell_line, lineage_num):
self.cell_line = cell_line
self.lineage_num = lineage_num
self.T = cell_line[0].T
self.t = cell_line[0].t
self.L = np.array([x.length[0][0] for x in cell_line])
self.spots = shared.get_parB_path(self.cell_line, self.T, self.lineage_num)
self.spot_storage = SpotStorage()
self.plot_setup()
self.plot_parB()
plt.show()
def get_parB_split(lin, lin_num):
""" Time until first ParB spot splits, or None if no split """
parB_paths = shared.get_parB_path(lin, lin[0].T, lin_num)
lin_t0 = lin[0].T[lin[0].frame - 1]
split_times = []
for path in parB_paths:
if type(path) is shared.TraceConnect:
if path.split_parent:
# it has split!
# get timing of split
split_times.append(path.timing[0] - lin_t0)
if not split_times:
return None
else:
return np.min(split_times)
def process():
lin_files = sorted(glob.glob("data/cell_lines/lineage*.npy"))
lineage_nums = sorted([int(re.search("lineage(\d+).npy", x).group(1)) for x in lin_files])
processable = []
traces = {}
for lineage_num in lineage_nums:
lineage_file = "data/cell_lines/lineage{0:02d}.npy".format(lineage_num)
cell_line = np.load(lineage_file)
T = cell_line[0].T
parB_paths = shared.get_parB_path(cell_line, T, lineage_num)
for spot_trace in parB_paths:
if hasattr(spot_trace, "spot_ids"):
traces[spot_trace.spot_ids[0]] = spot_trace
if type(spot_trace) is shared.TraceConnect and spot_trace.split_children:
processable.append((
spot_trace.split_children[0],
spot_trace.split_children[1],
spot_trace
))
velocities = []
growth_rates = []
elong_rates = []
for spot_pair in processable:
spot1 = traces[spot_pair[0]]
spot2 = traces[spot_pair[1]]
parent = spot_pair[2]
add_parent(parent, spot1)
add_parent(parent, spot2)
# take spot with minimum max timing as comparator (spot1)
if spot1.spots()["timing"].max() > spot2.spots()["timing"].max():
delta = get_delta(spot2, spot1)
else:
delta = get_delta(spot1, spot2)
if len(delta) >= 5:
interspot_velocity = get_velocity(delta)
if interspot_velocity:
velocities.append(interspot_velocity)
growth_rates.append(get_growth_rate(delta))
elong_rates.append(get_elong_rate(delta))
return velocities, growth_rates, elong_rates
def process():
lin_files = sorted(glob.glob("data/cell_lines/lineage*.npy"))
spots = []
for lf in lin_files:
lineage_num = int(re.search("lineage(\d+).npy", lf).group(1))
cell_line = np.load(lf)
elongation_rate = shared.get_elongation_rate(cell_line, discard=True)
if not elongation_rate or len(cell_line) < THRESHOLD_FRAMES:
continue
parB_paths = shared.get_parB_path(cell_line, cell_line[0].T, lineage_num)
spot_num = 1
for path in parB_paths:
spot_trace = path.spots()
lengths = list(path.len())
idx = 0
spot_data = {
"t": [],
"intensity": [],
"cell_length": lengths,
"x_mid": [],
"x_new": [],
"x_old": [],
"pole_known": cell_line[0].pole_assignment,
"spot_num": spot_num,
"lineage_num": lineage_num,
}
for x in spot_trace:
l = lengths[idx]
x_M = x[1]
x_N = x[1] + (l / 2)
x_O = l - x_N
spot_data["t"].append(x[0])
spot_data["intensity"].append(x[2])
spot_data["x_mid"].append(x_M)
spot_data["x_new"].append(x_N)
spot_data["x_old"].append(x_O)
idx += 1
spot_num += 1
if len(spot_data["t"]) >= THRESHOLD_FRAMES:
spots.append(spot_data)
# calculate diffusion parameters
d_mid, d_new, d_old = diffusion(spot_data)
s = pd.DataFrame(spots)
print(s)
def get_traces(orig_dir=None, two_spot=False, reuse=True):
data_hash = hashlib.sha1(os.getcwd().encode("utf8")).hexdigest()
if reuse and orig_dir and os.path.exists(os.path.join(orig_dir, "ParB_velocity", "data", data_hash)):
data_dir = os.path.join(orig_dir, "ParB_velocity", "data", data_hash)
files = sorted(glob.glob(os.path.join(data_dir, "*.pandas")))
spot_data = []
progress = progressbar.ProgressBar()
for f in progress(files):
spot_data.append(pd.read_pickle(f))
return spot_data
lin_files = sorted(glob.glob("data/cell_lines/lineage*.npy"))
lineage_nums = [int(re.search("lineage(\d+).npy", x).group(1)) for x in lin_files]
spot_data = []
progress = progressbar.ProgressBar()
for lineage_num, lf in progress(list(zip(lineage_nums, lin_files))):
cell_line = np.load(lf)
if not hasattr(cell_line[0], "pole_assignment") or cell_line[0].pole_assignment is None:
continue
# pole_assignment = cell_line[0].pole_assignment
T = cell_line[0].T
paths = shared.get_parB_path(cell_line, T, lineage_num)
if two_spot:
if len(paths) != 3:
continue
if len(cell_line[0].ParB) != 1:
continue
cell_elongation_rate = shared.get_elongation_rate(cell_line)
if cell_elongation_rate and cell_elongation_rate < 0:
cell_elongation_rate = 0
spot_num = 1
for path in paths:
# path.positions: distance from midcell
spot_trace = path.spots()
timing = []
d_mid = []
d_parA = []
intensity = []
lengths = path.len()
for x in spot_trace:
timing.append(x[0])
d_mid.append(x[1])
intensity.append(x[2])
c_idx = list(cell_line[0].t).index(x[0])
cell = cell_line[c_idx]
parA_mid = cell.ParA[0] - (cell.length[0][0] / 2)
dparA = np.abs(parA_mid - x[1])
d_parA.append(dparA)
data = pd.DataFrame(
data={
"timing": timing,
"d_mid": d_mid, # negative = closer to new pole
"d_parA": d_parA,
"intensity": intensity,
"cell_length": lengths,
},
)
data["d_new"] = data.d_mid + (data.cell_length / 2)
data["d_old"] = data.cell_length - data.d_new
path, subdir = os.path.split(os.getcwd())
topdir = os.path.basename(path)
data._path = os.getcwd()
data._top_dir = topdir
data._sub_dir = subdir
data._lineage_num = lineage_num
data._spot_num = spot_num
data._cell_line_id = cell_line[0].id
data._elongation_rate = cell_elongation_rate
data._hash = hashlib.sha256("{0}-{1}-{2}-{3}".format(
topdir,
subdir,
lineage_num,
spot_num,
).encode("utf-8")).hexdigest()
data._metadata = [
"_path", "_top_dir", "_sub_dir", "_lineage_num",
"_spot_num", "_cell_line_id",
"_elongation_rate", "_hash"
]
if orig_dir:
target_dir = os.path.join(orig_dir, "ParB_velocity", "data", data_hash)
if not os.path.exists(target_dir):
os.makedirs(target_dir)
data.to_pickle(os.path.join(
target_dir, "{0:03d}-{1:03d}.pandas".format(lineage_num, spot_num)
))
spot_data.append(data)
spot_num += 1
return spot_data
def plot_graphs(cell_line, lineage_num, num_plots=5, parA_heatmap=None, save=True, labels=None, um=False, plot_kwargs=None):
lineage = track.Lineage()
if save:
fig = plt.figure(figsize=(20, 10))
gs = matplotlib.gridspec.GridSpec(2, 3)
fig.patch.set_alpha(0)
spots_ParA = []
traces_ParA = []
L = []
T = shared.get_timings()
for cell in cell_line:
spots_ParA.append(cell.ParA)
traces_ParA.append(cell.parA_fluorescence_smoothed[::-1])
if um:
L.append(cell.length[0][0] * PX)
else:
L.append(cell.length[0][0])
L = np.array(L)
traces_ParA = np.concatenate(traces_ParA)
max_parA_intensity = traces_ParA.max()
start = cell_line[0].frame - 1
end = cell_line[-1].frame - 1
t = np.array(T[start:end + 1])
if save:
parA_heatmap = fig.add_subplot(gs[0, 0])
_despine(parA_heatmap)
# use Rectangles to generate a heatmap
i = 0
cmapper = plt.cm.get_cmap("afmhot")
for cell in cell_line:
trace = cell.parA_fluorescence_smoothed
if um:
l = cell.length[0][0] * PX
else:
l = cell.length[0][0]
x0 = t[i] - 8
y0 = -(l / 2)
increment = l / len(trace)
colours = cmapper(trace / max_parA_intensity)
i2 = 0
for _t in trace:
if _t < 0:
_t = 0
r = matplotlib.patches.Rectangle(
(x0, y0),
width=15,
height=increment,
facecolor=colours[i2],
edgecolor="none",
)
parA_heatmap.add_patch(r)
y0 += increment
i2 += 1
i += 1
if save:
parA_heatmap.set_ylabel(r"Distance from mid-cell (px)")
parA_heatmap.set_xlabel(r"Time (min)")
parA_heatmap.set_title("ParA")
poledict = poles.PoleAssign(lineage.frames).assign_poles()
if poledict[cell_line[0].id] is None:
parA_heatmap.plot(t, L / 2, "r-", lw=2)
parA_heatmap.plot(t, -(L / 2), "r-", lw=2)
else:
parA_heatmap.plot(t, L / 2, "k-", lw=2)
parA_heatmap.plot(t, -(L / 2), "k-", lw=2)
parAs = {}
i = 0
for s in spots_ParA:
midcell = s[2] / 2
spot = s[0] - midcell
parAs[t[i]] = spot
i += 1
parAs = np.array(sorted(parAs.items()))
if num_plots >= 2:
parA_heatmap.plot(
parAs[:, 0], parAs[:, 1],
lw=2, marker=".", mec="k", ms=10
)
decorate_daughters(cell_line, lineage, parA_heatmap, labels=labels, um=um)
parA_heatmap.patch.set_alpha(0)
if num_plots >= 3:
ax_parA = fig.add_subplot(gs[1, 0])
_despine(ax_parA)
ax_parA.set_title("ParA maximum")
ax_parA.plot(
parAs[:, 0], (L / 2) - parAs[:, 1],
marker=".", lw=2, mec="k", ms=10,
label="Distance from top pole"
)
ax_parA.plot(
parAs[:, 0], (L / 2) + parAs[:, 1],
marker=".", lw=2, mec="k", ms=10,
label="Distance from bottom pole"
)
ax_parA.set_xlabel(r"Time (min)")
ax_parA.set_ylabel(r"Distance (px)")
ax_parA.patch.set_alpha(0)
if num_plots >= 2:
parB_midcell = fig.add_subplot(gs[0, 1])
_despine(parB_midcell)
parB_midcell.set_title("ParB")
parB_midcell.plot(t, L / 2, "k-", lw=2, label="Cell poles")
parB_midcell.plot(t, -(L / 2), "k-", lw=2)
spots_ParB = shared.get_parB_path(cell_line, T, lineage_num)
spotnum = 1
for x in spots_ParB:
s = x.spots(False)
if hasattr(x, "split_parent") and x.split_parent:
for y in spots_ParB:
if y.spot_ids[-1] == x.split_parent:
textra = y.timing[-1:]
timings = np.concatenate([textra, s["timing"]])
posextra = np.array(y.position[-1:])
if um:
positions = np.concatenate([posextra * PX, s["position"] * PX])
else:
positions = np.concatenate([posextra, s["position"]])
break
else:
timings = s["timing"]
if um:
positions = s["position"] * PX
else:
positions = s["position"]
if num_plots >= 2:
ax_target = parB_midcell
label = "Spot {0}".format(spotnum)
else:
ax_target = parA_heatmap
label = "ParB"
if plot_kwargs:
ax_target.plot(
timings, positions, **plot_kwargs
)
else:
# ax_target.plot(
# timings, positions,
# lw=2,
# marker=".",
# markeredgecolor="k",
# ms=10,
# label=label,
# color=colour,
# )
ax_target.plot(
timings, positions,
lw=2,
marker=".",
markeredgecolor="k",
ms=10,
label=label,
color="k",
path_effects=[
matplotlib.patheffects.Stroke(
linewidth=2, foreground=(0.7, 0.7, 0.7)
)
],
)
dparA = []
for spot in s:
parApos = parAs[np.where(parAs[:, 0] == spot[0])][0, 1]
dpA = spot[1] - parApos
dparA.append(dpA)
x.parA_d = dparA
x.parA_dmean = np.mean(dparA)
x.parA_dsem = np.std(dparA) / np.sqrt(len(dparA))
x.spotnum = spotnum
x.intensity_mean = s["intensity"].mean()
spotnum += 1
if num_plots >= 2:
lines = parB_midcell.lines[::-1]
for l in lines:
parB_midcell.lines.remove(l)
parB_midcell.add_line(l)
parB_midcell.set_ylabel(r"Distance from mid-cell (px)")
parB_midcell.set_xlabel(r"Time (min)")
decorate_daughters(cell_line, lineage, parB_midcell, pad=5, labels=labels, um=um)
parB_midcell.legend(bbox_to_anchor=(1.35, 1))
parB_midcell.patch.set_alpha(0)
else:
if save:
parA_heatmap.legend(
[parA_heatmap.lines[-1]], [parA_heatmap.lines[-1].get_label()],
bbox_to_anchor=(1.2, 1)
)
if num_plots >= 4:
filtered = [x for x in spots_ParB if len(x) > 4]
if len(filtered) > 0:
ax_parB_closest = fig.add_subplot(gs[1, 1], sharex=ax_parA, sharey=ax_parA)
dmin = min(
filtered,
key=lambda x: np.abs(x.parA_dmean)
)
ax_parB_closest.set_title("ParB spot {0} (dmin)".format(dmin.spotnum))
_despine(ax_parB_closest)
s = dmin.spots(False)
dpol_t = (dmin.len() / 2) - s["position"]
dpol_b = (dmin.len() / 2) + s["position"]
ax_parB_closest.plot(
s["timing"], dpol_t,
marker=".", lw=2, mec="k", ms=10,
label="Distance from top pole"
)
ax_parB_closest.plot(
s["timing"], dpol_b,
marker=".", lw=2, mec="k", ms=10,
label="Distance from bottom pole"
)
ax_parB_closest.plot(
s["timing"], dmin.parA_d,
marker=".", lw=2, ms=10,
label="Distance from ParA focus"
)
ax_parB_closest.plot(
ax_parB_closest.get_xlim(),
[0, 0],
"k--"
)
ax_parB_closest.patch.set_alpha(0)
if num_plots >= 5:
imax = max(filtered, key=lambda x: x.intensity_mean)
if imax.id != dmin.id:
ax_parB_highest = fig.add_subplot(gs[1, 2], sharex=ax_parB_closest, sharey=ax_parA)
s = imax.spots(False)
dpol_t = (imax.len() / 2) - s["position"]
dpol_b = (imax.len() / 2) + s["position"]
ax_parB_highest.plot(s["timing"], dpol_t, marker=".", lw=2, mec="k", ms=10, label="Distance from top pole")
ax_parB_highest.plot(s["timing"], dpol_b, marker=".", lw=2, mec="k", ms=10, label="Distance from bottom pole")
ax_parB_highest.plot(s["timing"], imax.parA_d, marker=".", lw=2, mec="k", ms=10, label="Distance from ParA focus")
ax_parB_highest.plot(ax_parB_highest.get_xlim(), [0, 0], "k--")
_despine(ax_parB_highest)
ax_parB_highest.set_title("ParB Spot {0} (imax)".format(imax.spotnum))
ax_parB_highest.set_ylabel(r"Distance (px)")
ax_parB_highest.set_xlabel("Time (min)")
ax_parB_highest.patch.set_alpha(0)
ax_parB_highest.legend(bbox_to_anchor=(0.8, 1.35))
else:
ax_parB_closest.legend(bbox_to_anchor=(1.65, 1))
else:
ax_parB_closest.legend(bbox_to_anchor=(1.65, 1))
else:
ax_parA.legend(bbox_to_anchor=(1.65, 1))
if save:
plt.tight_layout()
fn = "data/data-lineage{0:02d}.pdf".format(lineage_num)
plt.savefig(fn)
print("Saved data file to {0}".format(fn))
def plot_graphs_parB_only(cell_line, lineage_num, ax_parB=None, save=True, labels=None, um=False):
if um:
L = np.array([x.length[0][0] * PX for x in cell_line])
else:
L = np.array([x.length[0][0] for x in cell_line])
T = shared.get_timings()
t = np.array(T[cell_line[0].frame - 1:cell_line[-1].frame])
lineage = track.Lineage()
if not ax_parB:
fig = plt.figure(figsize=(20 / 3, 5))
ax_parB = fig.add_subplot(111)
_despine(ax_parB)
ax_parB.set_title("ParB")
ax_parB.set_ylabel(r"Distance from mid-cell (px)")
ax_parB.set_xlabel(r"Time (min)")
poledict = poles.PoleAssign(lineage.frames).assign_poles()
if poledict[cell_line[0].id] is None:
ax_parB.plot(t, L / 2, "r-", lw=2, label="Cell poles")
ax_parB.plot(t, -(L / 2), "r-", lw=2)
else:
ax_parB.plot(t, L / 2, "k-", lw=2, label="Cell poles")
ax_parB.plot(t, -(L / 2), "k-", lw=2)
spots_ParB = shared.get_parB_path(cell_line, T, lineage_num)
spotnum = 1
colourwheel = sns.color_palette(n_colors=len(spots_ParB))
for x in spots_ParB:
colour = colourwheel[spotnum - 1]
s = x.spots(False)
if hasattr(x, "split_parent") and x.split_parent:
for y in spots_ParB:
if y.spot_ids[-1] == x.split_parent:
textra = y.timing[-1:]
timings = np.concatenate([textra, s["timing"]])
posextra = np.array(y.position[-1:])
if um:
positions = np.concatenate([posextra * PX, s["position"] * PX])
else:
positions = np.concatenate([posextra, s["position"]])
break
else:
timings = s["timing"]
if um:
positions = s["position"] * PX
else:
positions = s["position"]
ax_parB.plot(
timings, positions,
lw=2, marker=".",
mec="k", ms=10,
label="Spot {0}".format(spotnum),
color=colour
)
spotnum += 1
lines = ax_parB.lines[::-1]
for l in lines:
ax_parB.lines.remove(l)
ax_parB.add_line(l)
decorate_daughters(cell_line, lineage, ax_parB, pad=5, labels=labels, um=um)
if save:
ax_parB.legend(bbox_to_anchor=(1.35, 1))
ax_parB.patch.set_alpha(0)
plt.tight_layout()
fn = "data/data-lineage{0:02d}.pdf".format(lineage_num)
plt.savefig(fn)
print("Saved data file to {0}".format(fn))
plt.close()
def determine_par_path(self, force=False):
self.spots = shared.get_parB_path(self.cell_line, self.T, self.lineage_num, force=force)
def gen_xl(cell_line, lineage_num):
lineage_num = int(lineage_num)
T = cell_line[0].T
t = cell_line[0].t
spots_ParA = [x.ParA for x in cell_line]
spots_ParB = shared.get_parB_path(cell_line, T, lineage_num)
excel_wb = xlwt.Workbook()
ws_parA = excel_wb.add_sheet("ParA")
ws_parB = excel_wb.add_sheet("ParB")
ws_parA.write(0, 0, "Time")
ws_parA.write(0, 1, "Distance from top pole")
ws_parA.write(0, 2, "Distance from midcell")
ws_parA.write(0, 3, "Cell length")
ws_parA.write(0, 4, "Spot intensity")
parAs = {}
i = 0
for pos, inten, celllen in spots_ParA:
midcell = celllen / 2
spot = pos - midcell
ws_parA.write(i + 1, 0, int(t[i]))
ws_parA.write(i + 1, 1, float(pos))
ws_parA.write(i + 1, 2, float(spot))
ws_parA.write(i + 1, 3, float(celllen))
ws_parA.write(i + 1, 4, float(inten))
parAs[t[i]] = spot
ws_parB.write(i + 1, 1, float(celllen))
i += 1
ws_parB.write(0, 0, "Time")
ws_parB.write(0, 1, "Cell length")
i = 1
time_dict = {}
for time_ in t:
ws_parB.write(i, 0, int(time_))
time_dict[float(time_)] = i
i += 1
rmax = max(time_dict.values())
ws_parB.write(rmax + 2, 0, "Intensity mean:")
ws_parB.write(rmax + 3, 0, "Intensity SEM:")
ws_parB.write(rmax + 4, 0, "Distance from ParA (mean):")
ws_parB.write(rmax + 5, 0, "Distance from ParA (SEM):")
col = 2
spotnum = 1
for x in spots_ParB:
s = x.spots(False)
ws_parB.write(0, col, "Distance from mid-cell (Spot {0})".format(spotnum))
ws_parB.write(0, col + 1, "Intensity (Spot {0})".format(spotnum))
ws_parB.write(0, col + 2, "Distance from ParA (Spot {0})".format(spotnum))
parA_d = []
for spot in s:
r = time_dict[spot[0]]
ws_parB.write(r, col, float(spot[1]))
ws_parB.write(r, col + 1, float(spot[2]))
dpA = spot[1] - parAs[spot[0]]
parA_d.append(dpA)
ws_parB.write(r, col + 2, float(dpA))
# intensity mean and SEM for spot lineage
intensity_mean = s["intensity"].mean()
intensity_sem = s["intensity"].std() / np.sqrt(len(s["intensity"]))
ws_parB.write(rmax + 2, col + 1, float(intensity_mean))
ws_parB.write(rmax + 3, col + 1, float(intensity_sem))
# distance from ParA mean and SEM for spot lineage
parA_dmean = np.mean(parA_d)
parA_dsem = np.std(parA_d) / np.sqrt(len(parA_d))
ws_parB.write(rmax + 4, col + 2, float(parA_dmean))
ws_parB.write(rmax + 5, col + 2, float(parA_dsem))
col += 3
spotnum += 1
if not os.path.exists("data/xls"):
os.mkdir("data/xls")
fn = "data/xls/lineage{0:02d}.xls".format(lineage_num)
if os.path.exists(fn):
if not os.path.exists("data/xls/backups"):
os.mkdir("data/xls/backups")
backup_file = "data/xls/backups/{0}-lineage{1:02d}.xls".format(
datetime.datetime.strftime(datetime.datetime.now(), "%Y%m%d-%H%M"),
lineage_num
)
attempt = 2
while os.path.exists(backup_file):
first = backup_file.split(".xls")[0]
backup_file = "{0}.xls-{1}".format(first, attempt)
attempt += 1
print("Backing up previous xls file to {0}".format(backup_file))
os.rename(fn, backup_file)
print("Saving new xls file to {0}".format(fn))
excel_wb.save(fn)