Martin Chalfie takes the advice of a fellow teacher and applied to work in the laboratory of Jose Zadunaisky at Yale University where he studies chloride transport in the frog retina. His research leads to his first publication. With revived confidence, he returned to Harvard.

Still unsure about what to do after his college, Chalfie has a series of short-term jobs: he interviews people in hospitals to find out how they repair their electronic equipment for a Department of Education study, he substitutes his father selling dresses, he does some draft counselling, and sets up summer rock concerts. After he teaches for two years chemistry, first-year algebra, and an introductory social science course at Hamden Hall Country Day School in Hamden, Connecticut.

Chalfie returns to Harvard under Robert Perlman who has recently, with Pastan, revealed the role of cyclic AMP in catabolite repression in E. coli. At Harvard he studies catecholamine biosynthesis and secretion, particularly from the adrenal gland. Chalfie works in two other labs, those of Tom Wilson and Sue Leeman but he decides to do his final thesis in Perlman’s lab using cell suspensions from rat pheochromocytomas (adrenal tumors) to look at the biosynthesis and release of catecholamines.

Martin Chalfie receives with Osamu Shimomura and Roger Y. Tsien the Nobel Prize in Chemistry "for the discovery and development of the green fluorescent protein, GFP, one of the most important tools in bioscience."

Martin Chalfie joins the department of biological sciences at Columbia University and continues to study C. elegans touch mutants. He finds that some mutants are insensitive to touch because the cells are missing or incompletely differentiated, a result that leads to the study of how cell type is determined. In those years at the Columbia, precisely in 1898, Chalfie marries Tulle Hazelrigg.

Chalfie conducts his postdoctoral research at MRC Laboratory of Molecular Biology where he meets Sidney Brenner. Chalfie is interested in working on the genetics of touch and Brenner gives his approval. Chalfie familiarizes with various mutant phenotypes. He starts his first mutagenesis and begins looking for touch-insensitive mutants. He isolates touch mutants, works on microtubule structure, studies lineage mutants, investigates neural circuitry, and explores even on neurotransmitters.

Sam Ward offers Martin Chalfie a temporary place in his laboratory. Here Chalfie looks at C. elegans for the first time and gradually learns how to grow them. Chalfie attends the first international C. elegans meeting at Woods Hole where he acquires a new research project. Bob Horvitz encourages him to look at the work of John Sulston on touch-insensitive mutants and to continue that project.

Chalfie enters Harvard University. He begins to think of majoring in math, but soon decides to become instead a biochemistry’s major. For his senior thesis Chalfie works for Klaus Weber. He analyses the active site of the enzyme aspartate transcarbamylase by chemically modifying different amino acids within it. He fails all his experiments hence, demotivated, he spends his senior year taking non-science courses (except for one Physics course).

Martin Chalfie first hears about GFP in 1988 at a seminar given by Paul Brehm about bioluminescent organisms. Chalfie immediately becomes very excited about the idea of expressing the fluorescent protein in the nematode. In September 1992, after obtaining GFP DNA from Douglas Prasher, Chalfie asks his rotation student, Ghia Euskirchen to express GFP in E. coli. Following Euskirchen's successful expression in E. coli, Chalfie's technician Yuan Tu goes on to express GFP in C. elegans.

Martin Chalfie's first research experience is in the lab of Dr. Paul Kohn in the Biochemistry Department at the University of Illinois at Chicago between his sophomore and junior years. Kohn is interested in synthesizing furanosyl nucleosides (purines and pyrimidines linked to six carbon sugars with five member rings) as potential antitumor drugs. Chalfie has only few memories of this first laboratory experiences and not particularly enthusiastic.

Martin Chalfie is born in Chicago as the oldest of three sons of Vivian and Eli Chalfie. Since an early age he is interested in science. He loves to spend time with his first microscope and his chemistry set. He also enjoys reading.

Chalfie publishes what he considers his most influential paper, which describes the first use of the now-ubiquitous green fluorescent protein (GFP) as a reporter of gene expression. Since then GFP has become a fundamental tool of cell biology, developmental biology, genetics, neurobiology, and the medical sciences.