Isolation and analysis of genomic DNA clones encoding the third component of mouse complement

Area of Science:

• Genomics
• Molecular Biology
• Immunology

Background:

• The complement system is crucial for innate immunity.
• Mouse complement component 3 (C3) is a key protein in complement activation.
• Understanding C3 gene structure is vital for studying immune responses.

Purpose of the Study:

• To isolate and characterize genomic clones of the mouse C3 gene.
• To determine the physical size and organization of the C3 gene.
• To identify regulatory elements and predict protein features of C3.

Main Methods:

• Construction of a mouse genomic DNA library using phage lambda vector lambda 1059.
• Screening the library with a complementary DNA (cDNA) probe for C3.
• Isolation and characterization of recombinant phages containing C3 gene sequences.
• DNA sequencing to determine gene length and identify regulatory elements.

Main Results:

• Four distinct C3 genomic clones were isolated.
• Two clones contained overlapping inserts totaling 24 kilobase pairs, encompassing the complete C3 gene.
• The gene includes 5' and 3' flanking sequences, a T-A-T-A-A-A promoter element, and a poly(A) site.
• A 24-amino acid signal peptide was predicted at the N-terminus, encoded by the first exon.

Conclusions:

• The complete structure of the mouse C3 gene has been elucidated.
• The genomic organization provides insights into C3 gene regulation.
• The predicted signal peptide suggests a secretory pathway for C3.