Genome editing is a powerful tool for the analysis of targeted genes. Takuya
Tsubota and colleagues from the National Agriculture and Food Research
Organization in Japan previously found that a novel knock-in system, precise
integration into target chromosome (PITCh), allows the integration of a
donor vector with the hsp90 promoter and GFP into the silkworm biogenesis of
lysosome-related organelles complex 1, subunit 2 gene.

Then the team evaluated whether the same technique can be used for the
knock-in of other silkworm genes. The silkworm ku80 gene was selected as a
target and was efficiently mutated using a pair of transcription
activator-like effector nucleases (TALENs).

Microinjection of TALEN mRNAs mixed with the donor vector resulted in
significant expression of the GFP marker in G0 larvae. GFP expression was
detected in G1 individuals, suggesting that the integrated donor vector can
be inherited in the next generation. It was further found to be inherited in
the G2 generation.

The PITCh system offers a versatile tool for the knock-in of various genes,
and could contribute to the further promotion of sericultural studies.



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