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<i>Salmonella enterica</i> serovar Indiana (<i>S.</i> Indiana) is among the most prevalent serovars of <i>Salmonella</i> and is closely associated with foodborne diseases worldwide. In this study, we combined a recombinase polymerase amplification (RPA) technique with clustered regularly interspaced short palindromic repeat (CRISPR) and CRISPR-associated (Cas) protein Cas12b (CRISPR/Cas12b)-based biosensing in a one-pot platform to develop a novel one-step identification method for <i>S.</i> Indiana infection diagnosis. The entire RPA-CRISPR/Cas12b reaction can be completed at 41 °C within 1 h without the need for specific instruments. The optimal concentrations of Cas12b and single-guide RNA (sgRNA) for the reaction were the same at 250 nM. The single-stranded DNA (ssDNA) reporter 8C-FQ (5′-/6-FAM/CCCCCCCC/BHQ1/-3′) presented the best performance in the reaction compared with the other reporters. The limit of detection (LoD) of the RPA-CRISPR/Cas12b assay was 14.4 copies per reaction. As for specificity, we successfully identified four <i>S.</i> Indiana strains among twenty-two <i>Salmonella</i> strains without any false-positive results, presenting 100% accuracy for <i>S.</i> Indiana, and no cross-reactions were observed in eight other pathogens. Moreover, a total of 109 chicken carcasses were classified by the <i>S.</i> Indiana RPA-CRISPR assay and PCR methods from three processing points, including 43 post-shedding, 35 post-evisceration, and 31 post-chilling. There were 17 <i>S</i>. Indiana-positive samples identified during the whole processing step, consisting of nine post-shedding, five post-evisceration, and three post-chilling. The corresponding <i>S.</i> Indiana-positive rates of post-shedding, post-evisceration, and post-chilling were 20.93% (9/43), 14.29% (5/35), and 9.68% (3/31), respectively. Results from the <i>S.</i> Indiana one-step RPA-CRISPR/Cas12b assay were totally in agreement with those obtained using a traditional culture method, demonstrating 100% agreement with no false-positive or false-negative results observed. Altogether, the RPA-CRISPR/Cas12b assay developed in this study represents a promising, accurate, and simple diagnostic tool for <i>S.</i> Indiana detection.