After obtaining cDNA, double strand was synthesized.
Primers (degenerate primers) are designed according to the gene sequence of homologous species.
The target fragment is amplified by PCR (nested PCR can be used to increase specificity)
Rubber cutting, recycling and purification
Plasmid vector connection transfection
Sequencing of recombinant obtained by screening
After obtaining the gene sequence, the primers were designed again according to the sequence.
Run a 3-foot and 5-foot race
Re-sequencing confirmed the full-length gene.
Functional analysis of Arabidopsis mutant gene
This is the process of doctoral thesis experiment done by one of my classmates.
What we are doing is homologous cloning of a tree species.