In order to research the failure characteristics of segmental lining structure for Nanjing Yangtze River tunnel, a prototype test was carried out by using a “multi-function shield tunnel structure test system”. Water pressure and earth pressure were applied separately upon the prototype segmental lining structure in the test. And the failure characteristics of underwater shield tunnels under straight or staggered assembling were discussed respectively. The research results show that the damages of Nanjing Yangtze River tunnel are resulted mainly from bend cracking. Bend crack developing is always accompanied by the remarkable development of structure deformation and segment joint opening. The features of segment joint opening, cracking and deformation developing are different to the two different types of assembling structures. Compared with a straight assembling segmental structure, a staggered assembling segmental structure has a small deformation and crack width but fast cracking speed