Molecular Biology Resources


Links to protocols, programs & guides for molecular biology, e.g., sequence analysis, tree building, TRFLP analysis, available on the web.

Links for Sequence Analysis & Tree Building


Geneious
This is a program for all OS (Windows, Linux and Mac) for sequence analysis, etc. I have been using the Pro version for a few weeks and so far I am really happy with it. I like the sequence viewing, the contig assembly and the Blast features. The program allows you to batch BLAST your sequences against the NCBI database from within the program. You can tell it how many results you want to get back and they are delivered back to the program. VERY, VERY useful!!

FastGroupII
FastGroupII is a web-based tool for analyzing sequence data. It is specifically designed for dealing with large SSU rRNA sequence clone libraries. It contains several different algorithms for trimming sequences and grouping phylotypes based on sequence similarity. Also a very useful program for clustering sequences into OTUs and for downstream analysis. 

greengenes
Site specific for aligning and working with ribosomal sequences. The align function is excellent for aligning sequences to nearest neighbors and incorporates rRNA secondary structure in the algorithm.

UniFrac
This is an excellent program for analyzing microbial community sequence data. It is free online and very easy to use.

Fast UniFrac
Fast UniFrac is a new version of UniFrac that is specifically designed to handle very large datasets. Like UniFrac, Fast UniFrac provides a suite of tools for the comparison of microbial communities using phylogenetic information.

BioEdit
This is a really nice (free) program for sequence manipulation. Unfortunately it is only available for PC.

TreeView
I find that TreeView is a very nice program for viewing trees and exporting them as images. It is available for both Mac and PC but there are few options for text formatting and no options for illustrations.

TreeDyn
I just heard about this program and have not tried it out. It looks promising.

PAUP* 4.0
I am not an expert at PAUP and have had difficulty in remembering commands, etc. for building phylogenetic trees. Here are a few links that I found useful when battling through PAUP:

4Peaks
4Peaks is a program for visualize and edit their DNA sequence files on Mac OS X. I downloaded this program but have not been using it very much. It seems to be useful but not for large data sets. Brought to you by the same programmers of Papers.

eBio Tools
Another program for sequence analysis in Mac OS X. This is a nice program but I now default to Geneious for most of my sequence analysis.

Links for Terminal Restriction Fragment Length Polymorphism (TRFLP) Analysis


PAT-TRFLP: Phylogenetic Assignment Tool for TRFLP
PAT-TRFLP is a web-based tool for analyzing of TRFLP profiles of microbial communities. This is a very nice application that is easy to use! I like the fact that it is available on the web and able to be used by all OS users. Another bonus for this program: the user can define the database for matching profiles.

FragSort
The TRFLP Fragment Sorter Suite is a Windows based program for identifying peaks from TRFLP profiles. This program is excellent although there are a few stumbling points for getting it running for a user-specified database. Once up and running it is easily used. Two great features: numerous algorithms for incorporating fragment sizing error and uses a user-defined database for matching peaks.

Links for Denaturing Gradient Gel Electrophoresis (DGGE)


DGGE Help
This is a blog maintained by Dr. Stefan Green. The DGGE Help website contains guides and discussions for using and troubleshooting DGGE techniques.
Stefan Green's website: http://www.stefangreen.com/

Alternative Electrophoresis Buffer to Replace TAE or TBE (originally posted by Stefan J. Green on January 13, 2008 at http://ddgehelp.blogspot.com/).
I would like to advocate the use of three new electrophoresis buffers. Links to the relevant manuscripts are below. Also found below is a link to the company “Faster-better-media” selling these buffers.

(1) SB Buffer – for standard applications - 10 mM sodium boric acid (1X working solution is either 5 mM disodium borate decahydrate or 10 mM NaOH adjusted to pH 8.5 with boric acid; up to 50X concentrates can be made. 20X concentrate should have a pH of 8.0 → for more details see the manuscript below)
(2) LA Buffer – for longer DNA fragments (>3 kb) - 5 mM lithium acetate
(3) LB Buffer – for separation of small DNA fragments - 1 mM lithium boric acid (lithium hydroxide pH buffered to pH ~8.5 with boric acid)

The advantages of these buffers:
(a) they generate much less heat than TAE or TBE buffers
(b) they can be run at much high voltages (>300V in some cases) without fear of gels melting
(c) they can be much cheaper than TAE or TBE
(d) they do not interfere with standard applications such as band excision
(e) they last longer before needing to be replaced.

Faster-Better-Media website

References:
  • Brody JR, Kern SE. Sodium boric acid: A tris-less, cooler conductive medium for DNA electrophoresis. BioTechniques 2004; 36:214-216
  • Brody JR, Kern SE. History and principles of conductive media for standard DNA electrophoresis. Analytical Biochemistry 2004; 333:1-13.
  • Brody JR, Kern SE. Ultra-fast high-resolution agarose-electrophoresis of DNA and RNA using low-molarity conductive media. BioTechniques 2004; 37:598-602.