Tag: Rstats

Multiplex: Small multiple artwork from GPX tracks

I’d seen the small multiple artwork of running and cycling routes from Marcus Volz’s R package Strava all over the web. Ads for “posters of your GPS tracks” pop up on Reddit and I’d notice a few #Rstats people put up their posters on Twitter. I’ve had the package bookmarked for a while and this week I finally got round to generating a small multiple poster of some of my cycling routes.

I was pleased with the result and wanted to post it here. But also, running the code was not straightforward as I’ll explain below. If you want to generate your own plot read on.

The idea behind the poster is really nice. You get a kind of generative art-style poster. It looks nice and you can identify individual routes which is fun to do.

The instructions on the GitHub page are absolutely correct and the code should run out-of-the-box. The idea is that you download your Strava data and then make your plots. Unfortunately, it seems that a change in Strava’s data export policy (possibly related to GDPR changes) has broken the package. I found that there are two problems. First, Strava’s “download your data” link gives you a mix of formats (in my case GPX and FIT files), the package only works with GPX. Second, if there is any elevation data missing from a track, the data frame that is needed to make the poster is not built properly.

Going GPX only: In my case, I don’t keep all my data in Strava and instead use a local repository managed with RubiTrack. This software allows me to filter for the tracks I want and export them in GPX format. The only problem is that it generates one huge file with all the tracks enclosed. This gets read by the package as a single track. To fix this, I split the file using awk.

awk '/<trk>/{close(file);n++;}{file="track"n".gpx";print >> file;}' untitled.gpx

I could then discard track1.gpx which just had the xml header and then use the directory of gpx files.

The elevation problem: this affected only some of the tracks, so in the end the R code needed to be modified. The elevation data is not needed to make the posters so the file process_data.R needs editing, line 28 can be commented out and then line 32 should read:

result <- data.frame(lat = lat, lon = lon, time = time, type = type) %<%

This issue is raised on GitHub and has been closed, but the code doesn’t work with elevation blanks. If you run into this problem, this is the way I found to fix it. The other plots in the package which do use elevation will not run, but at least the poster can be made.

I exported the poster as PDF and then made some changes in Illustrator to give the result above.

—

The post title comes from Multiplex from Oliver’s Standing Stone LP from 1974.

By adventures in code, fun , , , , Leave a Comment

Pledging My Time III

I’ve previously crunched times for local Half and Full Marathons here on quantixed. Last weekend was the Kenilworth Half Marathon (2018) over a new course. I thought I’d have a look at the distributions of times and paces of the runners. The times are available here. If the Time and Category for finishers are saved as a csv, the script below works to generate the following plots.

Aggregated stats for the race are here. The beeswarm plot nicely shows the distribution of runners times and paces per category. There’s a bimodality to some of the age groups which is interesting. You can see from the average times that people get slower as they get older, as expected.

There was a roughly 2:1 split of M:F runners with a similar proportion in all categories. The ratio is similar for DNSers. The winning times were Andrew Savery of Leamington C A & C in MV35 with 01:12:51 and Polly Keen of Nuneaton Harriers in F sen with 01:23:46.

Congrats to everyone who ran and thanks to the organisers and all the supporters out on the course.


require(ggplot2)
require(ggbeeswarm)
file_name <- file.choose()
df1 <- read.csv(file_name, header = TRUE, stringsAsFactors = FALSE)
# aggregate M and F to a new category called Gender
df1$Gender <- ifelse(startsWith(df1$Category,"F"),"F","M")
# format Date column to POSIXct
df1$Time <- as.POSIXct(strptime(df1$Time, format = "%H:%M:%S"))
orig_var <- as.POSIXct("00:00:00", format = "%H:%M:%S")
p1 <- ggplot( data = df1, aes(x = Category,y = Time, color = Category)) + 
  geom_quasirandom(alpha = 0.5, stroke = 0) +
  stat_summary(fun.y = mean, geom = "point", size=2, aes(group = 1)) +
  scale_y_datetime(date_labels = "%H:%M:%S", limits = c(orig_var,NA))
p1
# instead of finishing time, let's look at pace (min/km)
df1$Pace <- as.numeric(difftime(df1$Time, orig_var) / 21.1) * 3600
df1$Pace <- as.POSIXct(df1$Pace, origin = orig_var, format = "%H:%M:%S")
p2 <- ggplot( data = df1, aes(x = Category,y = Pace, color = Category)) + 
  geom_quasirandom(alpha = 0.5, stroke = 0) +
  stat_summary(fun.y = mean, geom = "point", size=2, aes(group = 1)) +
  scale_y_datetime(date_labels = "%M:%S", limits = c(orig_var,NA))
p2
# calculate speeds rather than pace
df1$Speed <- 21.1 / as.numeric(difftime(df1$Time, orig_var))
p3 <- ggplot( data = df1, aes(x = Category, y = Speed, color = Category)) + 
  geom_quasirandom(alpha = 0.5, stroke = 0) +
  stat_summary(fun.y = mean, geom = "point", size=2, aes(group = 1)) +
  ylim(0,NA) + ylab("Speed (km/h)")
p3
# now make the same plots but by Gender rather than Category
p4 <- ggplot( data = df1, aes(x = Gender,y = Time, color = Gender)) + 
  geom_quasirandom(alpha = 0.5, stroke = 0) +
  stat_summary(fun.y = mean, geom = "point", size=2, aes(group = 1)) +
  scale_y_datetime(date_labels = "%H:%M:%S", limits = c(orig_var,NA))
p4
p5 <- ggplot( data = df1, aes(x = Gender,y = Pace, color = Gender)) + 
  geom_quasirandom(alpha = 0.5, stroke = 0) +
  stat_summary(fun.y = mean, geom = "point", size=2, aes(group = 1)) +
  scale_y_datetime(date_labels = "%M:%S", limits = c(orig_var,NA))
p5
p6 <- ggplot( data = df1, aes(x = Gender, y = Speed, color = Gender)) + 
  geom_quasirandom(alpha = 0.5, stroke = 0) +
  stat_summary(fun.y = mean, geom = "point", size=2, aes(group = 1)) +
  ylim(0,NA) + ylab("Speed (km/h)")
p6
ggsave("raceTimeByCat.png", plot = p1)
ggsave("racePaceByCat.png", plot = p2)
ggsave("raceSpeedByCat.png", plot = p3)
ggsave("raceTimeByGen.png", plot = p4)
ggsave("racePaceByGen.png", plot = p5)
ggsave("raceSpeedByGen.png", plot = p6)

Edit 2018-09-12T18:52:43Z I wasn’t happy with the plots and added a few more lines to look at gender as well as category. And show speed as well as pace and finishing time.

—

The post title is taken from “Pledging My Time” a track from Blonde on Blonde by Bob Dylan

By fun , One Comment

Rollercoaster III: yet more on Google Scholar

In a previous post I made a little R script to crunch Google Scholar data for a given scientist. The graphics were done in base R and looked a bit ropey. I thought I’d give the code a spring clean – it’s available here. The script is called ggScholar.R (rather than gScholar.R). Feel free to run it and raise an issue or leave a comment if you have some ideas.

I’m still learning how to get things looking how I want them using ggplot2, but this is an improvement on the base R version.

—

As described earlier I have many Rollercoaster songs in my library. This time it’s the song and album by slowcore/dream pop outfit Red House Painters.

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Ten Years vs The Spread: Calculating publication lag times in R

There have been several posts on this site about publication lag times. You can read them here. Lag times are the delays in the dissemination of scientific data introduced by the process of publishing the paper in a journal. Nowadays, your paper can be online in a few hours using a preprint server. However, this work is not peer reviewed. Journals organise a formal peer review and provide some sort of certification of the work. They typeset the work and all of this adds delays the dissemination of work in a journal.

To look at publication delays, you can use PubMed data, which is incomplete but can give insight into how long these delays can be. Previous posts have involved the use of a ruby script to make a csv file from PubMed XML output and then use this in Igor to calculate the publication lag times. There is another method detailed in this excellent post by Daniel Himmelstein.

I recently posted a figure for Nature Communications lag times on Twitter and was asked to generate others. I figured that I should write an R script and people can make their own!

The PubMedLagR code is available here with instructions for use.

A query for Nature Communications data at PubMed, such as:

nat commun[ta] AND 2000 : 2018[pdat] AND journal article[pt]

Retrieves all paper for this journal. The range from 2010 to 2018 is for illustration, this journal has only been in operation for these years. Filtering for journal articles rather and attempting to get rid of reviews and front matter is wise, but doesn’t always work. Again this journal doesn’t carry this material so this is for illustration. Getting your query right is very important.

Save the results in XML format and then run the R script as directed. This should give a csv of the data and a png of the lag times.

This is data from Nature Communications. Colleagues had two separate papers accepted at this journal and experienced long delays. I was interested to see if papers were generally taking longer to publish here. Of course we do not know why. Delays are partly the fault of the authors, the reviewers and the journal and it is not possible to say why publication lag times are increasing for this journal year-on-year. The journal has grown in terms of number of papers published, has this introduced inefficiencies? Are reviewers being slow to review? Are they being more demanding? Are Editors not marshalling the referee reports and providing clear guidance to authors? Allowing too much time and too many rounds of revision? Are authors being too slow to do further experimental work? The answer will be yes to some of these questions for some of the papers.

This is not to focus on Nature Communications, it’s one of a few journals that many colleagues complain is too slow to publish their work. With this code you can have a look at the journal you are interested in submitting to and consider whether there is a more rapid venue for your work.

Update:

I changed the code slightly and prettified the plots just a little. Below are some plots for Nature Cell Biology, Nature Neuroscience. I also did a search for clathrin or CRISPR papers over the same time period. These keyword searches are fairly flat, whereas the journal-specific increase in publication lag time can be seen.

The lag times at Nature Neuroscience look artificially low and then seem to have jumped up in 2016 to be something similar to Nature Cell Biology or Nature Communications.

Edit

I neglected to point out that the code truncates the y-axis in the bottom right plot to 1000 days or the maximum lag time, whichever is smaller. This is because it gets difficult to see the data points if there is an outlier, which might be due to an error in PubMed data.

A reader commented on Twitter that some poor paper had a lag time almost 1000 days. Well, due to the y-axis truncation we don’t see that 9 papers in Nature Communications since 2010 have lag times (RecAcc) of > 1000 days. The record holder has a lag time of 1561 days! I checked that this was not a PubMed error by looking at the dates on the paper.

Notes

Date information is not available in PubMed for every paper unfortunately. This is especially true of older papers.

The date information is supplied to PubMed from the journal. These dates are not necessarily accurate: 1) you can see occasional errors in the data, 2) journals sometimes “reset the clock” on papers and treat resubmissions as new submissions.

—

The post title is taken from “10 Years vs The Spread” by Wing-Tipped Sloat from the LP Chewyfoot. Obviously the song has nothing to do with smoothed kernel density estimates of journal publication lag times, but the title was incredibly apt.

By computing, publishing , , , 3 Comments

Rollercoaster II: more on Google Scholar citations

I’ve previously written about Google Scholar. Its usefulness and its instability. I just read a post by Jon Tennant on how to harvest Google Scholar data in R and I thought I would use his code as the basis to generate some nice plots based on Google Scholar data.

A script for R is below and can be found here. Graphics are base R but do the job.

First of all I took it for a spin on my own data. The outputs are shown here:

These were the most interesting plots that sprang to mind. First is a ranked citation plot which also shows y=x to find the Hirsch number. Second, was to look at total citations per year to all papers over time. Google Scholar shows the last few years of this plot in the profile page. Third, older papers accrue more citations, but how does this look for all papers? Finally, a prediction of what my H-index will do over time (no prizes for guessing that it will go up!). As Jon noted, the calculation comes from this paper.

While that’s interesting, we need to get  the data of a scholar with a huge number of papers and citations. Here is George Church.

At the time of writing he has 763 papers with over 90,000 citations in total and a H-index of 147. Interestingly ~10% of his total citations come from a monster paper in PNAS with Wally Gilbert in the mid 80s on genome sequencing.

Feel free to grab/fork this code and have a play yourself. If you have other ideas for plots or calculations, add a comment here or an issue at GitHub.

if(!require(scholar)){
     install.packages("scholar")
}
library(scholar)
# Add Google Scholar ID of interest here
ID <- ""
# If you didn't add one to the script prompt user to add one
if(ID == ""){
     ID <- readline(prompt="Enter Scholar ID: ")
}
# Get the citation history
citeByYear<-get_citation_history(ID)
# Get profile information
profile <- get_profile(ID)
# Get publications and save as a csv
pubs <- get_publications(ID)
write.csv(pubs, file = "citations.csv")
# Predict h-index
hIndex <- predict_h_index(ID)
# Now make some plots
# Plot of total citations by year
png(file = "citationsByYear.png")
plot(citeByYear$year,citeByYear$cites,
     type="h", xlab="Year", ylab = "Total Cites")
dev.off()
# Plot of ranked paper by citation with h
png(file = "citationsAndH.png")
plot(pubs$cites, type="l",
     xlab="Paper rank", ylab = "Citations per paper")
abline(0,1)
text(nrow(pubs),max(pubs$cites, na.rm = TRUE),
     profile$h_index)
dev.off()
# Plot of cites to paper by year
png(file = "citesByYear.png")
plot(pubs$year, pubs$cites,
     xlab="Year", ylab = "Citations per paper")
dev.off()
# Plot of h-index prediction
thisYear <- as.integer(format(Sys.Date(), "%Y"))
png(file = "hPred.png")
     plot(hIndex$years_ahead+thisYear,hIndex$h_index,
     ylim = c(0, max(hIndex$h_index, na.rm = TRUE)),
     type = "h",
     xlab="Year", ylab = "H-index prediction") 
dev.off()

Note that my previous code used a python script to grab Google Scholar data. While that script worked well, the scholar package for R seems a lot more reliable.

—

I have a surprising number of tracks in my library with Rollercoaster in the title. This time I will go with the Jesus & Mary Chain track from Honey’s Dead.

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Turn That Heartbeat Over Again: comparing wrist and chest-strap HRM

As a geek, the added bonus of exercise is the fun that you can have with the data you’ve generated. A recent conversation on Twitter about the accuracy of wrist-based HRMs got me thinking… how does a wrist-based HRM compare with a traditional chest-strap HRM? Conventional wisdom says that the chest-strap is more accurate, but my own experience of chest-strap HRMs is that they are a bit unreliable. Time to put it to the test.

I have a Garmin Fēnix 5 which records wrist-based HR and I have a Garmin chest-strap which uses ANT+ to transmit. I could pick up the ANT+ signal with a Garmin Edge 800 so that I could record both datasets simultaneously, on the same bike ride. Both the Fēnix and Edge can record GPS and Time (obviously) allowing accurate registration of the data. I also set both devices to receive cadence data via ANT+ from the same cadence/speed sensor so that I could control for (or at least look at) variability in recordings. I rode for a ~1 hr ~32 km to capture enough data. Obviously this is just one trial but the data gives a feel for the accuracy of the two HRMs. Biking, I figured was a fair activity since upper body and wrist movement is minimal, meaning that the contacts for both HRMs are more likely to stay in place than if I was running.

I’ll get to heart rate last. First, you can see that the GPS recording is virtually identical between the two units – so that’s a good sign. Second, elevation is pretty similar too. There’s a bit of divergence at the beginning and end of the ride, since those parts are over the same stretch of road, neither device looks totally accurate. I’d be inclined to trust the Fēnix here since has a newer altimeter in it. Third, cadence looks accurate. Remember this data is coming off the same sensor and so any divergence is in how it’s being logged. Finally, heart rate. You can see that at the beginning of the ride, both wrist-based and chest-strap HRMs struggle. Obviously I have no reference here to know which trace is correct, but the chest-strap recording looks like it has an erroneous low reading for several minutes but is otherwise normal. The wrist-based HRM looks like it is reading 120ish bpm values from the start and then snaps into reading the right thing after a while. The chest-strap makes best contact when some perspiration has built up, which might explain its readings. The comparisons are shown below in grey. The correlation is OK but not great. Compared to cadence, the data diverge a lot more which rules out simple logging differences as a cause.

I found a different story in Smart recording mode on the Fēnix. This is a lower frequency recording mode, which is recommended to preserve battery life for long activities.

So what can we see here? Well, the data from the Fēnix are more patchy but even so, the data are pretty similar except for heart rate. The Fēnix performs badly here. Again, you can see the drop out of the chest strap HRM for a few minutes at the start, but otherwise it seems pretty reliable.  The comparison graph for heart rate shows how poorly the wrist-based HRM measures heart rate, in this mode.

OK, this is just two rides, for one person – not exactly conclusive but it gives some idea about the data that are captured with each HRM.

Conclusions

Wrist-based HRM is pretty good (at the higher sampling rate only) especially considering how the technology works, plus chest-strap HRMs can be uncomfortable to wear, so wrist-based HRM may be all you need. If you are training to heart rate zones, or want the best data, chest-strap HRM is more reliable than wrist-based HRM generally. Neither are very good for very short activities (<15 min).

For nerds only

Comparisons like these are quite easy to do in desktop packages like Rubitrack (which I love) or Ascent or others. They tend to mask things like missing data points, they smooth out the data and getting the plots the way you want is not straightforward. So, I took the original FIT files from each unit and used these for processing. There’s a great package for R called cycleRtools. This worked great except for the smart recording data which was sampled irregularly and it turns out and the package requires monotonic sampling. I generated a gpx file and parsed the data for this activity in R using XML. I found this snippet on the web (modified slightly).

library(XML)
library(plyr)
filename <- "myfile.gpx"
gpx.raw <- xmlTreeParse(filename, useInternalNodes = TRUE)
rootNode <- xmlRoot(gpx.raw)
gpx.rawlist <- xmlToList(rootNode)$trk
gpx.list <- unlist(gpx.rawlist[names(gpx.rawlist) == "trkseg"], recursive = FALSE)
gpx <- do.call(rbind.fill, lapply(gpx.list, function(x) as.data.frame(t(unlist(x)), stringsAsFactors=F)))
names(gpx) <- c("ele", "time", "temp", "hr", "cad", "lat", "lon")

Otherwise:

library(cycleRtools)
edge <- as.data.frame(read_fit(file = file.choose(), format = TRUE, CP = NULL, sRPE = NULL))
write.csv(edge, file = "edge.csv", row.names = F)

The resulting data frames could be saved out as csv and read into Igor to make the plots. I wrote a quick function in Igor to resample all datasets at 1 Hz. The plots and layout were generated by hand.

—

The post title comes from “Turn That Heartbeat Over Again” by Steely Dan from Can’t Buy A Thrill

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I’m not following you II: Twitter data and R

My activity on twitter revolves around four accounts.

I try to segregate what happens on each account, and there’s inevitably some overlap. But what about overlap in followers?

What lucky people are following all four? How many only see the individual accounts?

It’s quite easy to look at this in R.

So there are 36 lucky people (or bots!) following all four accounts. I was interested in the followers of the quantixed account since it seemed to me that it attracts people from a slightly different sphere. It looks like about one-third of quantixed followers only follow quantixed, about one-third follow clathrin also and more or less the remainder are “all in” following three accounts or all four. CMCB followers are split about the same. The lab account is a bit different, with close to one-half of the followers also following clathrin.

Extra nerd points:

This is a Venn diagram and not an Euler plot. Venn just shows schematically the intersections and does not attempt to encode information in the area of each part. Euler plots for greater than three groups are hard to generate and to make any sense of what is shown. It is a dataviz problem to look at the proportions or lots of groups. A solution here would be to generate a further four Venn diagrams. On each, display the proportion for one category as a fraction or percentage

How to do it:

Last time, I described how to set up rtweet and make a Twitter app for use in R. You can use this to pull down lists of followers and extract their data. Using the intersect function you can work out the numbers of followers at each intersection. For four accounts, there will be 1 group of four, 4 groups of three, 6 groups of two. The VennDiagram package just needs the total numbers for all four groups and then details of the intersections, i.e. you don’t need to work out the groups minus their intersections – it does this for you.

library(rtweet)
library(httpuv)
library(VennDiagram)
## whatever name you assigned to your created app
appname <- "whatever_name"
## api key (example below is not a real key)
key <- "blah614h"
## api secret (example below is not a real key)
secret <- "blah614h"
## create token named "twitter_token"
twitter_token <- create_token(
app = appname,
consumer_key = key,
consumer_secret = secret)
clathrin_followers <- get_followers("clathrin", n = "all")
clathrin_followers_names <- lookup_users(clathrin_followers)
quantixed_followers <- get_followers("quantixed", n = "all")
quantixed_followers_names <- lookup_users(quantixed_followers)
cmcb_followers <- get_followers("Warwick_CMCB", n = "all")
cmcb_followers_names <- lookup_users(cmcb_followers)
roylelab_followers <- get_followers("roylelab", n = "all")
roylelab_followers_names <- lookup_users(roylelab_followers)
# a = clathrin
# b = quantixed
# c = cmcb
# d = roylelab
## now work out intersections
anb <- intersect(clathrin_followers_names$user_id,quantixed_followers_names$user_id)
anc <- intersect(clathrin_followers_names$user_id,cmcb_followers_names$user_id)
and <- intersect(clathrin_followers_names$user_id,roylelab_followers_names$user_id)
bnc <- intersect(quantixed_followers_names$user_id,cmcb_followers_names$user_id)
bnd <- intersect(quantixed_followers_names$user_id,roylelab_followers_names$user_id)
cnd <- intersect(cmcb_followers_names$user_id,roylelab_followers_names$user_id)
anbnc <- intersect(anb,cmcb_followers_names$user_id)
anbnd <- intersect(anb,roylelab_followers_names$user_id)
ancnd <- intersect(anc,roylelab_followers_names$user_id)
bncnd <- intersect(bnc,roylelab_followers_names$user_id)
anbncnd <- intersect(anbnc,roylelab_followers_names$user_id)
## four-set Venn diagram
venn.plot <- draw.quad.venn(
area1 = nrow(clathrin_followers_names),
area2 = nrow(quantixed_followers_names),
area3 = nrow(cmcb_followers_names),
area4 = nrow(roylelab_followers_names),
n12 = length(anb),
n13 = length(anc),
n14 = length(and),
n23 = length(bnc),
n24 = length(bnd),
n34 = length(cnd),
n123 = length(anbnc),
n124 = length(anbnd),
n134 = length(ancnd),
n234 = length(bncnd),
n1234 = length(anbncnd),
category = c("Clathrin", "quantixed", "CMCB", "RoyleLab"),
fill = c("dodgerblue1", "red", "goldenrod1", "green"),
lty = "dashed",
cex = 2,
cat.cex = 1.5,
cat.col = c("dodgerblue1", "red", "goldenrod1", "green"),
fontfamily = "Helvetica",
cat.fontfamily = "Helvetica"
);
# write to file
png(filename = "Quad_Venn_diagram.png");
grid.draw(venn.plot);
dev.off()

I’ll probably return to rtweet in future and will recycle the title if I do.

—

Like last time, the post title is from “I’m Not Following You” the final track from the 1997 LP of the same name from Edwyn Collins

By computing, fun , , 2 Comments

Frankly, Mr. Shankly

I read about Antonio Sánchez Chinchón’s clever approach to use the Travelling Salesperson algorithm to generate some math-art in R. The follow up was even nicer in my opinion, Pencil Scribbles. The subject was Boris Karloff as the monster in Frankenstein. I was interested in running the code (available here and here), so I thought I’d run it on a famous scientist.

By happy chance one of the most famous scientists of the 20th Century, Rosalind Franklin, shares a nominative prefix with the original subject. There is also a famous portrait of her that I thought would work well.

I first needed needed to clear up the background because it was too dark.

Now to run the TSP code.

The pencil scribbles version is nicer I think.

The R scripts basically ran out-of-the-box. I was using a new computer that didn’t have X11quartz on it nor the packages required, but once that they were installed I just needed to edit the line to use a local file in my working directory. The code just ran. The outputs FrankyTSP and Franky_scribbles didn’t even need to be renamed, given my subject’s name.

Thanks to Antonio for making the code available and so easy to use.

—

The post title comes from “Frankly, Mr. Shankly” by The Smiths which appears on The Queen is Dead. If the choice of post title needs an explanation, it wasn’t a good choice…

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Parallel lines: new paper on modelling mitotic microtubules in 3D

We have a new paper out! You can access it here.

The people

This paper really was a team effort. Faye Nixon and Tom Honnor are joint-first authors. Faye did most of the experimental work in the final months of her PhD and Tom came up with the idea for the mathematical modelling and helped to rewrite our analysis method in R. Other people helped in lots of ways. George did extra segmentation, rendering and movie making. Nick helped during the revisions of the paper. Ali helped to image samples… the list is quite long.

The paper in a nutshell

We used a 3D imaging technique called SBF-SEM to see microtubules in dividing cells, then used computers to describe their organisation.

What’s SBF-SEM?

Serial block face scanning electron microscopy. This method allows us to take an image of a cell and then remove a tiny slice, take another image and so on. We then have a pile of images which covers the entire cell. Next we need to put them back together and make some sense of them.

How do you do that?

We use a computer to track where all the microtubules are in the cell. In dividing cells – in mitosis – the microtubules are in the form of a mitotic spindle. This is a machine that the cell builds to share the chromosomes to the two new cells. It’s very important that this process goes right. If it fails, mistakes can lead to diseases such as cancer. Before we started, it wasn’t known whether SBF-SEM had the power to see microtubules, but we show in this paper that it is possible.

We can see lots of other cool things inside the cell too like chromosomes, kinetochores, mitochondria, membranes. We made many interesting observations in the paper, although the focus was on the microtubules.

So you can see all the microtubules, what’s interesting about that?

The interesting thing is that our resolution is really good, and is at a large scale. This means we can determine the direction of all the microtubules in the spindle and use this for understanding how well the microtubules are organised. Previous work had suggested that proteins whose expression is altered in cancer cause changes in the organisation of spindle microtubules. Our computational methods allowed us to test these ideas for the first time.

Resolution at a large scale, what does that mean?

The spindle is made of thousands of microtubules. With a normal light microscope, we can see the spindle but we can’t tell individual microtubules apart. There are improvements in light microscopy (called super-resolution) but even with those improvements, right in the body of the spindle it is still not possible to resolve individual microtubules. SBF-SEM can do this. It doesn’t have the best resolution available though. A method called Electron Tomography has much higher resolution. However, to image microtubules at this large scale (meaning for one whole spindle), it would take months or years of effort! SBF-SEM takes a few hours. Our resolution is better than light microscopy, worse than electron tomography, but because we can see the whole spindle and image more samples, it has huge benefits.

What mathematical modelling did you do?

Cells are beautiful things but they are far from perfect. The microtubules in a mitotic spindle follow a pattern, but don’t do so exactly. So what we did was to create a “virtual spindle” where each microtubule had been made perfect. It was a bit like “photoshopping” the cell. Instead of straightening the noses of actresses, we corrected the path of every microtubule. How much photoshopping was needed told us how imperfect the microtubule’s direction was. This measure – which was a readout of microtubule “wonkiness” – could be done on thousands of microtubules and tell us whether cancer-associated proteins really cause the microtubules to lose organisation.

The publication process

The paper is published in Journal of Cell Science and it was a great experience. Last November, we put up a preprint on this work and left it up for a few weeks. We got some great feedback and modified the paper a bit before submitting it to a journal. One reviewer gave us a long list of useful comments that we needed to address. However, the other two reviewers didn’t think our paper was a big enough breakthrough for that journal. Our paper was rejected*. This can happen sometimes and it is frustrating as an author because it is difficult for anybody to judge which papers will go on to make an impact and which ones won’t. One of the two reviewers thought that because the resolution of SBF-SEM is lower than electron tomography, our paper was not good enough. The other one thought that because SBF-SEM will not surpass light microscopy as an imaging method (really!**) and because EM cannot be done live (the cells have to be fixed), it was not enough of a breakthrough. As I explained above, the power is that SBF-SEM is between these two methods. Somehow, the referees weren’t convinced. We did some more work, revised the paper, and sent it to J Cell Sci.

J Cell Sci is a great journal which is published by Company of Biologists, a not-for-profit organisation who put a lot of money back into cell biology in the UK. They are preprint friendly, they allow the submission of papers in any format, and most importantly, they have a fast-track*** option. This allowed me to send on the reviews we had and including our response to them. They sent the paper back to the reviewer who had a list of useful comments and they were happy with the changes we made. It was accepted just 18 days after we sent it in and it was online 8 days later. I’m really pleased with the whole publishing experience with J Cell Sci.

 

* I’m writing about this because we all have papers rejected. There’s no shame in that at all. Moreover, it’s obvious from the dates on the preprint and on the JCS paper that our manuscript was rejected from another journal first.

** Anyone who knows something about microscopy will find this amusing and/or ridiculous.

*** Fast-track is offered by lots of journals nowadays. It allows authors to send in a paper that has been reviewed elsewhere with the peer review file. How the paper has been revised in light of those comments is assessed by at the Editor and one peer reviewer.

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Parallel lines is of course the title of the seminal Blondie LP. I have used this title before for a blog post, but it matches the topic so well.

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The International Language of Screaming

A couple of recent projects have meant that I had to get to grips more seriously with R and with MATLAB. Regular readers will know that I am a die-hard IgorPro user. Trying to tackle a new IDE is a frustrating experience, as anyone who has tried to speak a foreign language will know. The speed with which you can do stuff (or get your point across) is very slow. Not only that, but… if you could just revert to your mother tongue it would be so much easier…

The Babel Fish
The Babel Fish from http://hitchhikers.wikia.com/wiki/Babel_Fish

What I needed was something like a Babel Fish. As I’m sure you’ll know, this fish is the creation of Douglas Adams. It allows instant translation of any language. The only downside is that you have to insert the fish into your ear.

The closest thing to the Babel Fish in computing is the cheat sheet. These sheets are typically a huge list of basic commands that you’ll need as you get going. I found a nice page which had cheat sheets which allowed easy interchange between R, MATLAB and python. There was no Igor version. Luckily, a user on IgorExchange had taken the R and MATLAB page and added some Igor commands. This was good, but it was a bit rough and incomplete. I took this version, formatted it for GitHub flavored markdown, and made some edits.

The repo is here. I hope it’s useful for others. I learned a lot putting it together. If you are an experienced user of R, MATLAB or IGOR (or better still can speak one or more of these languages), please fork and make edits or suggest changes via GitHub issues, or by leaving a comment on this page if you are not into GitHub. Thanks!

R-MATLAB-IGOR-CheatSheet

Here is a little snapshot to whet your appetite. Bon appetit!

cssnapshot

 

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The post title is taken from “The International Language of Screaming” by Super Furry Animals from their Radiator LP. Released as a single, the flip-side had a version called NoK which featured the backing tracking to the single. Gruff sings the welsh alphabet with no letter K.

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