Uniform amplification of low-input DNA is important for applications across biology,including single-cell genomics,forensic science,and microbial and viral sequencing.However,the requisite biochemical amplification me...Uniform amplification of low-input DNA is important for applications across biology,including single-cell genomics,forensic science,and microbial and viral sequencing.However,the requisite biochemical amplification methods are prone to bias,skewing sequence proportions and obscuring signals relating to copy number.Digital droplet multiple displacement amplification enables uniform amplification but requires expert knowledge of microfluidics to generate monodisperse emulsions.In addition,existing microfluidic methods are tedious and labor intensive for preparing many samples.Here,we introduce rapid-emulsification multiple displacement amplification,a method to generate monodisperse droplets with a hand-held syringe and hierarchical droplet splitter.Although conventional microfluidic devices require >10 min to emulsify a sample,our system requires tens of seconds and yields data of equivalent quality.We demonstrate the approach by using it to accurately measure copy number variation(CNV)in single cancer cells.展开更多
基金We thank Angus Sidore and Freeman Lan for helpful scientific discussions.This work was supported by the UCSF Division of Hematology-Oncology Perkins Philanthropy(PLP)the National Science Foundation CAREER Award(Grant Number DBI-1253293)+2 种基金the National Institutes of Health(NIH)(Grant Numbers HG007233-01,R01-EB019453-01,and DP2-AR068129-01)the Defense Advanced Research Projects Agency Living Foundries Program(Contract Numbers HR0011-12-C-0065,N66001-12-C-4211,and HR0011-12-C-0066)Fold F(x)Program(Contract Number DE-AC02-05CH11231).
文摘Uniform amplification of low-input DNA is important for applications across biology,including single-cell genomics,forensic science,and microbial and viral sequencing.However,the requisite biochemical amplification methods are prone to bias,skewing sequence proportions and obscuring signals relating to copy number.Digital droplet multiple displacement amplification enables uniform amplification but requires expert knowledge of microfluidics to generate monodisperse emulsions.In addition,existing microfluidic methods are tedious and labor intensive for preparing many samples.Here,we introduce rapid-emulsification multiple displacement amplification,a method to generate monodisperse droplets with a hand-held syringe and hierarchical droplet splitter.Although conventional microfluidic devices require >10 min to emulsify a sample,our system requires tens of seconds and yields data of equivalent quality.We demonstrate the approach by using it to accurately measure copy number variation(CNV)in single cancer cells.
