In the manuscript “Advances in genome editing technology and its promising application in evolutionary and ecological studies”, the authors did a briefly introduction of three programmable gene editing technology, including ZFN, TALEN and RGN, in model organism. Then they computationally identified the possible target sites for TALEN and RGNs in 26 non-model organisms. Finally they summarized the potential applications of TALEN and RGNs in genetic revolution and the function of domesticated genes. The idea and the big frame for this review is good and interesting, but unfortunately, because of the limited knowledge and lacking of expertise in genome engineering field, the authors made several mistakes in basic definition. Sometimes, the writing is misleading and self-contradict.

Major comments:

1. Gene targeting/modification is DNA level change in genome, while gene activation/repression is gene expression regulation at mRNA level. e.g. “Different functional experiments demand different gene modification types, such as gene over-expression, gene knockdown and gene targeting, which includes gene knockout and gene knockin.” “Gene knockdown and gene over-expression” are not “gene modification” as no DNA sequences change.

2. The authors are confused with CRISPR VS CRISPR/Cas9 nuclease, also TALE/TALE VS TALEN. It’s misleading and inappropriate. e.g. a. In the abstract, “ZFN (Zinc Finger Nuclease), TALEN (Transcription Activator-Like Effector Nuclease), and CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) can trigger genome editing easily….” Here you should use “CRISPR/Cas9 nucleases”. “CRISPR” and “CRISPR/Cas9 nuclease” are totally different. “CRISPR” stands for “Clustered Regularly Interspaced Short Palindromic Repeats”, but not nuclease. b. “Similarly, TALEN could carry out regulation by custom-fusing the TALE DNA-binding domain with effector proteins, such as Mxi1 and VP64.” Here, you cannot use “TALEN”.

3. Self-contradict e.g. a. “ZFN (Zinc Finger Nuclease) [7, 8], TALEN (Transcription Activator-Like Effector Nuclease) [9, 10] and CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) [11, 12] systems now make it possible for scientists to modify the genome easily, efficiently and cheaply.” VS “ZFN has limited target sites because of the 3-nucleotides recognizing model, the system is also very expensive and difficult to assemble.” b. “ZFN (Zinc Finger Nuclease), TALEN (Transcription Activator-Like Effector Nuclease), and CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) can trigger genome editing easily, precisely and with no limitations by organism.” VS “These technologies have been successfully used in various model and non-model organisms with wide genome limitations and high efficiency.”

4. Mistakes e.g. a. “Another approach to improve CRISPR is to relieve recognition constraints of NGG PAM.” This is not correct at all. Stringent PAM sequences requirement is one of the major advantages of S. pyogenes Cas9 over other Cas proteins. b. In Table 1, 1) Applicable organism for site-specific recombinase technology is definitely not limited to mice and drosophila, at least including human cells and zebrafish as well. 2) CRISPR is not sensitive to methylation as reported in Hsu, P.D. et al, Nat Biotech.2013.

Minor comments:

Too many for each, just give a couple of examples:

1. Inappropriate reference: e.g.

2. Ref 10 is a ZFN paper, but the authors cited as a TALEN paper

3. Ref 13~17 only covers ZFN and TALEN paper, the authors cannot use them for ZFN, TALEN and CRISPR

4. Typo: e.g. The email address of corresponding author Wen Wang should be wwang@mail.kiz.ac.cn, but not wwang@mail,kiz.ac.cn

5. Grammar: e.g. “Different functional experiments demand different gene modification types, such as gene over-expression, gene knockdown and gene targeting, which includes gene knockout and gene knockin.” Should be “Different functional experiments demand different gene modification types, such as gene over-expression, gene knockdown and gene targeting which includes gene knockout and gene knockin.”

Level of interest An article of limited interest Quality of written English Not suitable for publication unless extensively edited Statistical review No, the manuscript does not need to be seen by a statistician. Declaration of competing interests I declare that I have no competing interests

The authors have addressed reviewers' critics in a reasonable fashion. Level of interest An article of limited interest

Quality of written English Acceptable Statistical review No, the manuscript does not need to be seen by a statistician.

Declaration of competing interests I declare that I have no competing interests