Monosynaptic viral tracers are essential tools for dissecting neuronal connectomes and for targeted delivery of molecular sensors and effectors. Viral toxicity and complex multi-injection protocols are major limiting application barriers. To overcome these barriers, we developed an anterograde monosynaptic H129_Amp tracer system based on HSV-1 strain H129. The H129_Amp tracer system consists of two components: an H129-dTK-T2-pac^Flox helper which assists H129_Amp tracer’s propagation and transneuronal monosynaptic transmission. The shared viral features of tracer/helper allow for simultaneous single-injection and subsequent high expression efficiency from multiple-copy of expression cassettes in H129_Amp tracer. These improvements of H129_Amp tracer system shorten experiment duration from 28-day to 5-day for fast-bright monosynaptic tracing. The lack of toxic viral genes in the H129_Amp tracer minimizes toxicity in postsynaptic neurons, thus offering the potential for functional anterograde mapping and long-term tracer delivery of genetic payloads. The H129_Amp tracer system is a powerful tracing tool for revealing neuronal connectomes.

单突触病毒示踪剂是解剖神经元连接体和靶向递送分子传感器和效应器的重要工具。病毒毒性和复杂的多次注射方案是主要的限制性应用障碍。为了克服这些障碍，我们开发了一种基于 HSV-1 菌株 H129的顺行单突触 H129 _{Amp示踪剂系统。}H129 _Amp示踪剂系统由两个组件组成：一个 H129-dTK-T2- pac ^Flox助手，它协助 H129 _Amp示踪剂的传播和跨神经元单突触传递。示踪剂/辅助病毒的共同病毒特征允许同时单次注射和随后从 H129 _{Amp中的多拷贝表达盒进行高表达效率}示踪剂。_{H129 Amp}示踪系统的这些改进将快亮单突触示踪的实验时间从 28 天缩短到 5 天。_{H129 Amp}示踪剂中缺乏毒性病毒基因可最大限度地减少突触后神经元的毒性，从而提供功能性顺行映射和遗传有效载荷的长期示踪剂传递的潜力。H129 _Amp示踪系统是一种强大的示踪工具，用于揭示神经元连接体。