In order to solve the problem of high concentration of antibiotic resistance genes (ARGs) in the secondary effluent of the sewage treatment plant, the molecular weight distribution and the main existence forms of ARGs in the secondary effluent was investigated in this study. And the effects of environmental factors such as the reaction time, temperature and pH value on the conjugation and transfer of plasmids were explored to provide research basis for reclaimed water treatment. The results show that the ARGs in the secondary effluent are mainly concentrated in the molecular weight range greater than 100k Da, and the concentration of free ARGs is higher than the concentration of cellular ARGs. The plasmid such as pHS-AVC-LW1144 could be achieved conjugation and transfer by the spreading among the same species or cross species which led to an increase of the plasmid concentration. The concentration of the plasmid showed an increasing trend during the reaction period of 0 to 48 h, and the maximum concentration was reached at the 48th hour; low temperature environment (4℃) and alkaline environment ( pH = 9) is more beneficial to the conjugation and transfer of resistant plasmids.