Screened and Pooled BAC library

We have been constructing high-quality cDNA and DNA libraries since 1997 and some of our clients include Agriculture Canada, Pioneer Hi-Bred Int'l (DuPont), Bayer, Cargill and the Max Planck Institutes in Germany. We guarantee a quick turnaround-time, unbeatable pricing and large inserts.

References are proudly provided.

We also offer BAC library construction in custom and shuttle vectors! Please contact areti@biost.com

POOLED BAC LIBRARY

A pooled BAC library is an invaluable tool for PCR-screening. It allows for the easy retrieval of BAC clones of interest. This type of library is especially useful when genome size is large and creating an arrayed library is cost-prohibitive.

 

Highlights

  1. We typically construct a minimum 5X BAC library which provides a 99.3% probability of finding the target sequence. Additional coverage and 2-enzyme digestion is available (additional fee). We pride ourselves in tailoring our services to suit our client’s needs.
  2. BAC clones are then pooled and distributed in 96-well plates. For example, a BAC library created from a 3Gb species will consist of at least 125,000 BAC clones (AIS of 125kb) distributed into 3 x 96-well plates. Each well will contain about 400-500 original BAC clones.
    1. Coverage is guaranteed. Amount of clones received is based on a guaranteed coverage and 120Kb average insert size.
  3. HMW DNA isolation and QC by HMW DNA enzymatic digestion are included.
  4. FREE and unlimited storage of Pooled BAC libraries at our facilities for future screening!
Related services

 

  1. Screening: The pooled BAC library can be PCR-screened using client-provided PCR primer pair(s) which specifically amplifies a segment in the target area. Price per screening is dependent on genome size. Client can decided the number of positive BAC clones he wishes to purchase.
  2. Row and Column Pools derived from Pooled BAC library: We can also provide the following pools which will allow client to immediately begin screening.
    1. Row pools from Superpool plates: isolated DNA.
    2. Column pools from Superpool plates: isolated DNA.
Advantages:
The pooled BAC library can be screened hundreds of times and stored indefinitely. Our pooling method keeps the library in a compact format which is very easy to store. Copies can be easily made and economically shipped to collaborators worldwide.
Drawback:
The only disadvantage to this type of library is that it cannot be used for physical mapping or full-genome sequencing

 

 

Our strengths

  1. We guarantee coverage.
  2. High average insert and even insert size distribution (see Graph 2 below).
  3. We perform HMW DNA extraction at our facilities.
  4. We have experience with plant, fungal, bacterial and animal libraries.
  5. We have extensive experience with GC-rich organisms and marine bacteria.
  6. Our arrayed BAC libraries are shipped in bar-coded microplates.
  7. We have extensive experience shipping worldwide.
  8. Shipment temperature of non-arrayed libraries is always monitored using a BLUE FLAG Temperature Data logger.
  9. We provide technical support.
  10. BAC libraries are the client's property and are never used internally, sold or distributed.
  11. We provide references
Table 1: A few libraries constructed in the past with obtained average insert sizes.

Starting material Insert size
Musmusculus 180 Kb
Zea mays 150 Kb
P. brassicacearum 160 Kb
P. omnivore 185 Kb
Arabidopsis thaliana 150Kb
Homo sapiens 215 Kb


Figure 1: PFG following NotI digestion of insect BAC clones ( Mamestra configurata ) with an average insert size of 178Kb

Insect


Figure 2: PFG following NotI digestion of fungal BAC clones P. omnivora) with an average insert size of 185Kb.

fungal


Graph 1: BAC clone insert size distribution (average of 140Kb) of a Soybean BAC library (Glycine max).


Quote request


Please email our Project Manager, Ms. Areti Karadimos, at areti@biost.com and include the following details. A quote will be provided in less than 24 hours.

  • - Name of organism of interest.
  • - Type of available starting material (leaves, cells, biopsy tissue, etc.)
  • - Is starting material limiting?
  • - What is the intended use for the BAC library (short-term and long-term if applicable).
  • - Known or estimated genome size of species of interest (haploid).
  • - Desired genomic coverage.

 

Importance of a BAC library


BAC libraries are essential elements in doing large-scale genome research such as genome sequencing, physical maps, and gene cloning. Bacterial artificial chromosomes (BAC) cloning technology was developed in early 1990s and quickly gained popularity. This technology was used in applications for genomic analysis that included large-scale physical mapping and genomic sequencing although this technology was developed much later than Yeast Artificial Chromosome (YAC) and cosmid cloning technologies. This should be attributed to the technical advantages of BAC cloning over YAC and cosmid cloning system summarized as follows:
 
  1. Bigger insert size (up to 300 kb) than cosmid and other plasmid system.
  2. Much more stable than yeast.
  3. Bacterial clones and libraries grow much faster than YAC.
  4. Easier in handling and gridding, DNA preparation.
  5. We have extensive experience with GC-rich organisms and marine bacteria.
  6. Our Allows for more efficient screening by either hybridization or PCR based screening.sive experience shipping worldwide.
The major applications of BAC library construction are in large-scale physical mapping and genomic sequencing and encompass the following applications:


 
Isolation of Gene clusters for entire metabolic pathway
A BAC library can be employed to scan for gene pathways in drug discovery, gene pathways in various diseases, plant genetics and microbial metabolic pathways. A variety of applications derive from construction of BAC libraries including BAC Macro arrays, gene isolation by pooled PCR, homology based hybridization using various DNA probes.

Prepare FISH probes
Fluorescence In Situ Hybridization (FISH) employs BAC vectors as probes to illumine and analyze tissue samples to understand chromosomal rearrangements, amplifications, deletions and aneusomies. Usually fluorescent probes for specific section(s) of a chromosome are used to label a locus of a chromosome within a cell sample where the specified genetic sequences are found.

Complementation for map-based gene cloning using Binary vector in plants
A BAC library can be used to interrogate the genetic structure of plant genomes as well as mutant genotype analysis using complementation techniques. This endows the scientist with a reliable tool to study global gene expression in plant species and to isolate individual genes.

Genomic walking
This allows direct genome sequencing using BAC libraries instead of sequencing directly from genomes, which is a very difficult proposition and allows study of gene structure using the entire genome.