Primer Design

Two sets of LAMP primers, which are targeted specific for IS6110 and IS1081, respectively, were designed using Primer Explorer V5 (http://primerexplorer.jp/e) according to the principles described by Notomi et al. (2000). Each primer set consisted of two inner primers (FIP, BIP), two outer primers (F3, B3) and two loop primers (LF, LB). To achieve the LFB detection, the FIP primer was labeled with FITC at the 5’ end, and was named as FIP*. The specificity of primers was confirmed by sequence alignment analysis using the basic local alignment search tool (BLAST). The locations and sequences of primers were shown in Figure 1 and Table 1 . Primers used in this study were synthesized by TianyiHuiyuan Biotechnology Co., Ltd (Beijing, China).

Primers specific to IS6110 and IS1081 of MTB used for LAMP-LFB assay. Sequence and locations of insertion sequences IS6110 and IS1081 used to design LAMP primers. The nucleotide sequences of IS6110 (A) and IS1081 (B) are shown. Primer FIP included F2 and F1c; Primer BIP included B1c and B2. The direction of arrows indicated the primer from 5’ to 3’.

The sequences of primer sets used in this study.

F3, Forward outer primer; B3, Backward outer primer; FIP, Forward inner primer; BIP, Backward inner primer; LF, Forward loop primer; LB, Backward loop primer; FIP*, Forward inner primer labeled with FITC at 5’ end.