The fat and amylose content of rice play a regulatory role in the biosynthesis of resistant starch subtype 5 （RS5）， which is the form of the RS in cooked rice. Investigating the genetic effects of the Waxy （Wx） gene， which regulates amylose biosynthesis， and the PAL6 gene involved in the fat synthesis， on different RS subtypes， as well as the relationship between rice quality and RS， is valuable for molecular breeding and quality improvement of rice with high RS. In this study， 69 rice germplasms with varying amylose contents were evaluated for RS content in cooked rice （RSc）， RS content in retrograded rice （RSr）， and cooking and eating quality traits. Functional variants of Wx and PAL6 were identified to assess their genetic effects and haplotype functions on RSc and RSr. Results indicated that both Wx and PAL6 significantly influence RS biosynthesis in rice， although the effect of PAL6 was comparatively less. The RS content were significantly higher （P<0.01） in Wx^a and Wx-T genotypes compared to Wx^b and Wx-C genotypes， while RSr and RSb（RSr-RSc）levels were significantly higher （P<0.05 or P<0.01） in PAL6-2 type than in PAL6-1 type. Haplotype analysis revealed that Hap1 （Wx^a-Wx-T-PAL6-1）， Hap2 （Wx^a-Wx-T-PAL6-2）， and Hap3 （Wx^a-Wx-C-PAL6-2） significantly increased RSc and RSr. Significant positive correlation （P<0.01） were observed between RSc and RSr on apparent amylose content， hot paste viscosity， cold paste viscosity， setback value， and consistence value. In contrast， significant negative correlations （P<0.05 or P<0.01） were found with gel consistency， breakdown value， pasting temperature， and peak viscosity. RSr was also negatively correlated with crude fat content， indicating that combination of fat and amylose during rice cooking might inhibit short-term retrogradation， thereby affecting RSr formation. Among the high RS haplotype germplasm， six with superior quality， CG133R， GBC120， GBC116， RILC-20， Shuhui 885 and RILC-15， were identified. This study revealed novel haplotypes associated with high RS content in rice， providing a theoretical foundation and germplasm resources for molecular breeding and quality improvement of rice with high RS.