The　determination　of　specie-specific　DNA　sequences　is　a　key　factor　for　identification　of　animal　species　and　detection　of　meat　adulteration.　Herein,　a　simple　homogeneous　electrochemical　biosensor　was　developed　for　sensitive　detection　of　specie-specific　DNA　sequences　from　meat　products　based　on　high　efficient　and　specific　dual-output　toehold-mediated　strand　displacement　(TMSD).　After　incubation　with　target　DNA,　large　amount　of　methylene　blue　(electrochemical　signal　molecule)　labeled　probes　(MB-P)　were　released　from　preformed　DNA　duplex　structures　by　the　process　of　dual-output　TMSD　amplification.　The　free　MB-P　could　be　further　digested　by　Exonuclease　I,　and　the　enzymatic　products　contain　little　negative　charge　could　diffuse　to　the　surface　of　indium　tin　oxide　electrode,　generating　significantly　electrochemical　signal.　As　a　result,　the　designed　biosensor　showed　a　broad　dynamic　range　from　0.01　pM　to　100　pM,　with　a　low　detection　limit　of　8.2　fM,　and　ideal　selectivity　and　reproducibility.　Meanwhile,　the　approach　exhibited　acceptable　accuracy　for　the　detection　of　specie-specific　DNA　sequences,　and　possessed　the　potential　application　for　the　identification　of　animal　species　from　meat　products.　©　2020　Elsevier　B.V.