Amaranthus palmeri was first reported in Turkey in 2016, and an immediate heavy infestation of the weed was found in fruit orchards and summer crops such as maize, cotton, and sunflower. There have been farmers' complaints about the ineffective control of Palmer amaranth through the use of glyphosate and some sulfonylureas herbicides. Hence, this study aimed to determine the possible herbicide resistance evolution in Palmer amaranth against glyphosate and acetolactate synthase (ALS) herbicides. Seeds of 21 Palmer amaranth populations were collected from five provinces of Turkey where control problems with glyphosate and ALS inhibitors were reported in maize fields. Seeds of certain biotypes categorized as resistant or susceptible were grown to obtain the F2 generation. A single‐dose experiment determined the possible resistance to ALS inhibitors and glyphosate among the 21 populations. Of this, 18 populations were included in the subsequent dose–response experiments due to evident survival. Based on ED50 values from the dose–response experiment, SNU‐04 and ADN‐21 biotypes had the highest resistance index for glyphosate which was more than 7. The biotypes ADN‐21, OSM‐15, and DIR‐09 recorded the highest ED50 value with a resistance index of 9.21–10.35 after nicosulfuron application. Whereas, the biotypes SNU‐04, OSM‐15, and ADN‐21 were with the highest ED50 value and resistance index of 6.41–7.44, after the application of foramsulfuron + iodosulfuron methyl‐sodium. The increase in genomic 5‐enolpyruvylshikimate‐3‐phosphate synthase (EPSPS) copy number has been observed in suspected cases that have been accepted as the molecular basis for the development of resistance against glyphosate. The sequence alignment results for the ALS gene contained Ala122Val and Pro197Arg mutations related to target‐site resistance against ALS herbicides.