Case Study 1—13.7 kb gene synthesis and plasmid modification

The 13.7 kb gene sequence was synthesized, assembled, and cloned into the designated vector site specified by the customer. The sequence was verified to be 100% correct by sequencing, and high-quality plasmids were successfully delivered by GenCefe.

Difficulties of the Project

• The sequence length reaches 13.7 kb, and it is prone to deletions during assembly

• The vector is provided by the customer, and there is no suitable restriction site for the insertion of the target gene on the vector

• Low copy vector, difficult to prepare

GenCefe Solutions

• The proposal was designed by experienced technical experts; using our proprietary assembly technology to ensure the correct full-length sequence

• Carried out vector modification, and successfully clone the target sequence into the designated position of the modified vector

• Self-developed competent cells and formula medium were used to increase the number of plasmid copies.

Case Study 2—difficult-to-synthesis gene sequence (PolyA-rich)

The gene sequence containing 110 consecutive PolyA has been successfully synthesized and verified by sequencing. And the 100% sequence correct plasmids were delivered to customers.

Difficulties of the Project

The sequence contains a large repeat structure, including a continuous PolyA structure of up to 110 bases. Such consecutive single-base repeats can lead to plasmid instability and abnormal sequencing signals.

GenCefe Solutions

• A specific synthetic scheme was designed for this sequence.

• Optimized construction and cloning protocols to reduce mutations.

• Optimized sequencing protocols to ensure accurate validation.

Case Study 3—difficult-to-synthesis gene sequence (GC-rich)

We have successfully synthesized and verified the gene sequence containing high GC contains, direct repeats,

 and polymer structures. And the

100% sequence correct plasmids were delivered to customers.

Difficulties of the Project

• The GC-rich region accounts for >35% of the total length, and the sequence GC content fluctuates notably. Such structure often leads to non-specific amplification and causes failure to amplify the target gene.

• Direct repeats plus polymer structures account for >30% of the total sequence. Repetitive sequences cause difficulties in assembly, while polymer structures cause sequencing difficulties.

GenCefe Solutions

• A specific synthesis scheme is designed for this sequence, and the gene sequence is synthesized in segments.

• Assembled using proprietary assembly technology to ensure the correct full-length sequence.

• Optimize construction and cloning protocols to reduce mutations.

• Optimize sequencing protocols to ensure accurate validation.

GenCefe Gene Synthesis Services