Fosmids

The BAC library was made from DNA from strain CC-503, the same strain sequenced by JGI. Fosmid clones were prepared as genomic DNA fragments in the fosmid vector pCCIFOS (Epicentre). Fosmids are similar to BACS (also available from the CRC) in that they contain large pieces of genomic DNA, but they offer several advantages for many genetic projects. First of all, there are more than 75,000 clones in the collection, and each region of the genome is covered in almost every case by multiple fosmids.  Fosmid inserts are shorter than BACs, averaging 35 – 40 kb, so they contain fewer genes.  It’s easy to isolate large quantities of fosmid DNA because the copy number of the fosmids in their bacterial hosts is greatly increased by treating cultures with arabinose. The construction and use of fosmid libraries at the JGI is described in this publication:

McNeal JR, Leebens-Mack JH, Arumuganathan K, Kuehl JV, Boore JL, DePamphilis CW. Using partial genomic fosmid libraries for sequencing complete organellar genomes. Biotechniques. 2006;41(1):69-73. doi:10.2144/000112202

Fosmid end sequences obtained by JGI are aligned with the genome sequence under JBrowse. For ver5.5 sequence, the fosmid sequences are displayed on the “BLAST-aligned BAC and fosmid end reads” track. The fosmid end sequence names are of the form “VTP[number]”. The two ends of the same fosmid clone have the same VTP name, but they differ in the extensions .b1, .b2, .g1 or .g2. The PTQ sequences are BAC end sequences.  Remember that you are ordering whole genomic clones not end sequences, so all you need to provide is the VTP number. To order, cut and paste the VTP numbers (e.g. VTP 12345) into the boxes below.

Protocol

From: Armstrong, Z; Rahfeld, P. Withers, SG. 2017. Methods Enzymol. 597:3-23.

Inoculate fosmid clones into 5 mL of TB media-containing 12.5 μg/mL chloramphenicol and 100 μg/mL arabinose. Incubate overnight at 37°C with shaking.

To order, enter the VTP number into the boxes below.