Abstract: A 90d feeding trial was conducted to determine the dietary lysine requirement of juvenile rice field eel (Monopterus albus), with an initial weight of (7.64±0.04) g reared in indoor aerated tanks. Six isonitrogenous and isoenergetic practical diets were formulated with graded levels of lysine (2.31%, 2.61%, 2.91%, 3.51%, 4.71%, and 7.11% dry matter). The result showed that: (1) The weight gain rate (WGR), specific growth rate (SGR), protein retention rate (PRR), and protein efficiency ratio (PER) of M. albus increased firstly and then decreased with the increase of dietary lysine level, and the feed conversion ratio (FCR) decreased first and then increased. Conversely, the FCR decreased initially and then increased. A dietary lysine level of 3.59% significantly improved the WGR, SGR, PRR, and PER and reduced FCR (P<0.05). A level of 3.51% significantly increased the relative expression of mTOR, S6K1, and IGF-1 in the liver, while decreasing the relative expression of eIF4E-BP2 (P<0.05). (2) With the increase of dietary lysine level, the crude protein in the muscle of juvenile M. albus increased firstly and then plateaued, while crude fat increased firstly and then decreased. The 7.11% lysine level significantly increased the contents of crude protein, total amino acid (TAA), essential amino acid (EAA), and non-essential amino acid (NEAA) in muscle (P<0.05). The 3.51% level significantly increased the crude fat content, EAA/TAA, and EAA/NEAA (P<0.05). (3) With the increase of dietary lysine level, the serum total protein (TP) and albumin (ALB) showed an increasing trend, and the 7.11% level was significantly higher than the 2.31% level (P<0.05). Serum glutamic oxalacetic transaminase activity (GOT) and serum ammonia (Sa) decreased firstly and then increased, while lysozyme (LZM) activity increased firstly and then decreased. Sa at 3.51% was significantly lower than that at other levels (P<0.05). GOT at 2.91% was significantly lower than that at 2.31% (P<0.05), and LZM at 4.71% was significantly higher than the level between 2.31% and 2.91% (P<0.05). (4) The activity of trypsin in hepatopancreas and foregut, as well as lipase activity in foregut, increased firstly and then decreased with increasing dietary lysine levels. Levels between 2.91% and 3.51% significantly increased trypsin and lipase activity in juvenile M. albus (P<0.05). In summary, the appropriate dietary lysine level can improve the digestive ability and non-specific immunity, promote the expression of GH/IGF and mTOR pathway related genes, and enhance the protein deposition, thereby improving the growth performance of juvenile M. albus. The optimal lysine requirement of M. albus (7.64±0.04 g) is estimated to be between 3.38% and 3.55% (6.57% to 6.90% of feed protein) based on a broken-line analysis of the relationships with WGR, SGR, FCR, PRR, and PER.