Construction of a fosmid library uses an improved cosmid plasmid vector. The vector system is constructed after introducing pBAC to PUCcos and fusing.

Features of the vector system:

• ŸDue to the insertion of the E. coli fertility factor (F-factor), it exists in the host bacteria as a single copy form and is very stable;
• The vector has an induced oriV high-copy, replication starting point. If needed, it can be induced to high copy (about 50);
• Ÿ The high randomness ensures equal frequency of every fragment of the DNA in the library;
• Ÿ The inserted fragments have high uniformity, generally about 40Kb.

Applications:

• Ÿ   Construction of whole-genome physical maps;
• Ÿ   Research of gene function and gene expression regulation;
• Ÿ   Map-based cloning of genes conferring important traits;
• Ÿ   Gene structure and function analysis.

Workflow:

Figure 1. Workflow of fosmid library construction

Bioinformatics Contents:

The main results of library qualification include storage capacity, insert segment size, homologous alignment, and randomness analysis.

Sample Requirements:

• ŸSample condition: Samples should be of DNA or fresh tissue. Plant samples should be etiolated seedlings.
• ŸSample concentration and purity: The concentration should be ≥ 50 ng/μL, the main band of genomic DNA ≥ 40 kb, the total DNA ≥ 50 μg, and OD_260/280 = 1.8 to 2.0. The DNA solutions should be free of visible contamination.

Turnaround Time:

The standard turnaround time for library construction using the workflow (above) is 30 working days.

Completion Milestone and Results Submission:

Completion Milestone:
The project is considered complete when the insert fragment size is approximately 40 kb, the empty clones rate is ≤ 5%, and the capacity of the library=genome size × coverage/insert fragment size.

Results submission:
Results include 96 or 384 bacterial preservation plates and a library inspection report.