Molecular Biology...From Scratch?

May222013

i-heart-histo:

A brief history of microscopy by i-heart-histo
c2000 BC
The Chinese use water microscopes made of a lens and a water filled tube to better visualize smaller objects.
1590
Hans Jansen and his son Zacharias Jansen invent the compound microscope.
1609
Galileo Galilei develops a compound microscope with a convex and concave lens. Calling it the occhiolino - the little eye.
1625
The term ‘microscope’ is coined by Giovanni Faber of Bamberg, an anology with the word ‘telescope’
1665
Robert Hooke publishes Micrographia and coins the word ‘cell’ after his examination of cork bark.
1674
Anton van Leuwenhoek develops the compound microscope to optimize it for observing biological specimens.
1860s
Ernst Abbe discovers the Abbe sine condition for manipulating the axis of optical systems to improving sharpess of images. This breakthrough in microscope design was exploited by microscope manufacturers Zeiss and Leitz resulting in a microscope boom.
1920
Olympus manufacture their first microscope - the Asahi.
1957
The Olympus DF Biological Microscope becomes the first microscope to feature an attached light source rather than a mirror that reflects light on the specimen.
1976
The popular CH series of Olympus microscopes appear in universities and colleges around the world. Chances are your college still uses these lab teaching scopes (or the slightly newer CH2 version).
1993
Introduction of a unique Y-shaped design for the microscope body for enhancing optics.
2004
Confocal and virtual microscopy are now common place.

I like the GIF

A brief history of microscopy by i-heart-histo

c2000 BC

The Chinese use water microscopes made of a lens and a water filled tube to better visualize smaller objects.

1590

Hans Jansen and his son Zacharias Jansen invent the compound microscope.

1609

Galileo Galilei develops a compound microscope with a convex and concave lens. Calling it the occhiolino - the little eye.

1625

The term ‘microscope’ is coined by Giovanni Faber of Bamberg, an anology with the word ‘telescope’

1665

Robert Hooke publishes Micrographia and coins the word ‘cell’ after his examination of cork bark.

1674

Anton van Leuwenhoek develops the compound microscope to optimize it for observing biological specimens.

1860s

Ernst Abbe discovers the Abbe sine condition for manipulating the axis of optical systems to improving sharpess of images. This breakthrough in microscope design was exploited by microscope manufacturers Zeiss and Leitz resulting in a microscope boom.

1920

Olympus manufacture their first microscope - the Asahi.

1957

The Olympus DF Biological Microscope becomes the first microscope to feature an attached light source rather than a mirror that reflects light on the specimen.

1976

The popular CH series of Olympus microscopes appear in universities and colleges around the world. Chances are your college still uses these lab teaching scopes (or the slightly newer CH2 version).

1993

Introduction of a unique Y-shaped design for the microscope body for enhancing optics.

2004

Confocal and virtual microscopy are now common place.

I like the GIF

(via molecularlifesciences)

May142013

Neurospora growing on a sugar cane stalk . Neurospora is efficient at using dead plant matter, >130 genes for hydrolyzing complex carbohydrates found in plant cell walls (Pectin, Cellulose, Chitin etc) are encoded in a genome with ~10,000 total genes although the plant pathogens Aspergillus and Fusarium have an even greater number. These enzymes are then secreted into the environment.

Science neurospora biology fungi mycology cell walls pathogens fusarium aspergillus cellulose

8PM

Tip of the day GATK Indel Calling

Remember to set the Genotype Likelihoods Model (-glm) to BOTH when running Unified Genotyper.

I knew that filtering/Variant Quality Score Recalibration have different procedures for SNPs/INDELs so I had left the INDEL step to last (bad idea). I kept finding 0 INDELs for my data set (when I had a huge number of SNP calls). Turns out I had forgotten to set the -glm to BOTH way back at the Unified Genotyper Step, whoops. I’ll probably go back and run on INDELs only later, but for now mystery solved.

GATK RNA-seq snp genomics bioinformatics science biology

May72013

biological-chemistry:

Multiple ribosomes bind to one transcript in prokaryotes.

Really cool

(via molecularlifesciences)

May62013

PLOS ONE: Development of Strategies for SNP Detection in RNA-Seq Data: Application to Lymphoblastoid Cell Lines and Evaluation Using 1000 Genomes Data

There has been some debate over whether RNA-Seq produces good enough SNP calling data in mammals This paper shows that >90% of the known SNPs could be detected in the RNA-Seq data set.

SNP RNA-Seq genomics GATK sequencing next generation sequencing

May32013

“The way we try to recruit girls into STEM fields is all wrong. We typically compare them to some great woman or someone that has gone before them. We are saying, “Hey, you can be like Madam Curie or Sally Ride.” It is recruiting by intimidation. We need to change that message. We need to recruit by appealing to WHY we need them in STEM. We NEED you to help make the world a better place We NEED you to help discover the cure for cancer. We NEED you because you have the ability to change the course of humanity for the better.”

Tim Holt on why we still see the number of females in STEM fields fall way behind their male counterparts. Also see how geography paved the way for women in science.

(↬ gender and science)

love it

(Source: explore-blog, via women-in-science)

May22013

VariantEval

VariantEval produces some great metrics using the CompOverlap module, I had alot of trouble discerning how to find comparisons between more than 2 VCF files (I have four conditions). This allows several files to be compared to a single comparison track ( I used a VCF that should represent background genotype). The module outputs metrics such as the number of Concordant loci, which I found very helpful to determine how SNPs might accumulate with the transfer process.

GATK SNP bioinformatics genome NGS RNA-Seq VCF

April222013

WHEN PEOPLE WAKE UP EARLY TO GO TO THE GYM BEFORE CLASS

whatshouldwecallgradschool:

April182013

CummeRbund - BioStar

Life saving tip for getting cummeRbund to create “myGenes” on a user defined list of genes as determined by gene name

R cummeRbund RNA-seq next generation sequencing biology science bioinformatics

April22013

Tophat and Cufflinks Example Command

I’m taking advantage of the new release of the Neurospora crassa genome and the new version of Cuffdiff to run my RNA-Seq data through its paces on the command line version of the Tuxedo suite. I’m going to post examples of the arguments I used in these programs here occasionally as there aren’t many examples and I may forget.

Tophat

tophat -G /path/to/GTF/file/ -o /path/to/output/directory/ -p 2 /path/to/index/directory/index_Base_Name/ /path/to/fastq/file/

-p is the argument for multi-threading which vastly speeds up the process of mapping, p can be set to any number less than or equal to the number of cores

Cufflinks

cufflinks -o /path/to/output/directory -p 2 -b /path/to/Reference.fa -u -N -G /path/to/GTF/file/ /path/to/accepted_hits.bam/

where -b is the argument for bias correction, -u is the multi-read correct and -N is the upper quartile normalization

In cuffmerge or cuffcompare remember to set the -s option to specify a reference genome to get all outputs

genomics tophat cufflinks science biology

6PM

WHEN THE UNDERGRAD TRIES TO CHANGE THE RADIO STATION

whatshouldwecallgradschool:

Funnier… when it gets left on what the they chose and I’m trying to figure out who i work with really loves recent Top 40.

6PM

A peril of snpEFF with a custom database

So you’ve decided to run snpEff with a custom reference genome and you finally figured how to use vi commands and build the database. If you see something like the image below where 0 protein coding genes are identified

This tells you that you need to follow the FAQ at snpEff for this condition, which means that you have treat every transcript as coding (default). However, the GATK recommends using the -onlyCoding option which only looks at the coding transcripts. If you use this option with a database that does not have protein coding info you do not get any snps annotated to the exons as below

Removing the -onlyCoding true argument results in SNPs being detected in exons as shown below

You’ll notice that my data does not have any SNPs detected in the introns/splice sites, this is expected as this is RNA-Seq data and I specifically limited the SNP calling to intervals in the transcript annotation file.

What’s important to note is this can be a problem with any reference/transcript file combo. I used the previous Neurospora build from Broad, and even though Broad is also responsible for GATK the genome did not play well with a tool that is part of the GATK pipeline.

A final note about snpEff output the genes.txt output reports values for all the genes in the transcript file even if the value is 0 (ala Cuffdiff). So you cannot go scanning this file with your favorite gene in mind without looking carefully at the output.

snpEff genomics science biology bioinformatics GATK SNP Neurospora

March262013

Yep Tophat, which I am running on command line for the first time can go to hell.

But on the other hand I’ve finally got a handle on GATK.

science bioinformatics Mac Linux Tophat

March252013

Starting Out in Bioinformatics, Unix and Perl for Biologists

If you’re not like me and you’d like to know what you’re doing before you start throwing arguments into the command line. Here is a primer on UNIX and Perl that was recommended on multiple Seqanswers threads. I may take a look at it when I have a spare day.

science biology bioinformatics grad school resources perl unix next generation sequencing

5PM

Students I taught in Intro Bio are now old enough to be sending me linkedin requests. Sigh.

I'm a PhD candidate where the motto seems to be "we're molecular biologists, we can do that!" and often on the cheap. I'll be discussing techniques, grad school and science. And maybe finding some humor in the mess.