Colorimetric nucleic acid detection on paper. microchip using loop mediated isothermal. amplification and crystal violet dye

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Hannah Osborne
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1 SUPPLEMENTARY INFORMATION Colorimetric nucleic acid detection on paper microchip using loop mediated isothermal amplification and crystal violet dye Sharmili Roy, Noor Faizah Mohd-Naim, Mohammadali Safavieh* and Minhaz Uddin Ahmed* Biosensors and Biotechnology Laboratory, Chemical Science Programme, Faculty of Science, Universiti Brunei Darussalam, Jalan Tungku Link, Gadong, BE 1410, Brunei Darussalam PAPRSB Institute of Health Science, Universiti Brunei Darussalam. Jalan Tungku Link, Gadong, BE 1410, Brunei Darussalam. Division of Engineering in Medicine, Brigham and Women s Hospital-Harvard Medical School, 75 Francis Street, Boston, Massachusetts 02115, United States

2 2 UV-vis spectral analysis with LAMP DNA UV-vis analysis was performed using LAMP amplicons including CV and LCV at different absorbances ranging from 400 to 700 nm at ph 7.5. Results obtained from UV-vis confirmed a significant difference in peak separation between that of LCV and LCV+LAMP DNA. The measurement of different concentrations of DNA were tested to verify the color change and its absorbance peak. The maximum color change and highest absorbance peak was obtained at a wavelength of 600 nm. 4.5nmol of CV solution was used in combination with sodium sulfite to make the solution colorless (in the form of LCV) with peak absorbance height of ~ 0.0. Upon addition of DNA, a peak absorbance height of ~ 0.3 was observed, marking a clear change in absorbance in the presence and absence of DNA. Figure S1. Absorbance spectroscopy of LCV dye without (red) and with DNA (blue). 2

3 3 Table S1: Analysis of the specificity of porcine primers with different raw meat samples. Sample and species name Detection of LAMP amplicons Normalized Relative intensity(%) Pork, Sus scrofa Yes Wild boar, Sus scrofa Yes Sheep, Ovis aries No Ostrich, Struthio camelus No Goat, Capra aegagrus phircus No Turkey, Meleagris gallopavo No Buffalo, Bison bison No Horse, Equus caballus No Duck, Anas platyrhynchos No

4 4 Table S2: Analysis of the specificity of porcine primers with different processed meat samples. Sample and species name Detection of LAMP amplicons Normalized Relative intensity (%) Spiced pork cubes, Sus scrofa Yes Pork mince with bean paste, Sus scrofa Yes Chopped pork & ham, Sus scrofa Yes Chao san si (pork and bamboo shoot), Sus Yes scrofa Mutton luncheon with chicken, Puffinus No tenuirostris and Gallus gallus Corned beef, Bos taurus No Chicken luncheon meat, Gallus gallus No 100 Beef loaf, Bos Taurus No Chicken luncheon meat, Gallus gallus No Mallow bakes, Bos taurus No Chamallow, Bos taurus No Marshmallows, Bos taurus No Boar meat, Sus scrofa Yes Corned mutton, Puffinus tenuirostris No Chicken luncheon meat, Gallus gallus No Curry beef, Bos taurus No

5 5 Chicken luncheon meat, Gallus gallus No Corned ostrich, Struthio camelus No Lamb curry with potatoes, Ovis aeris No Duck meat, Anas platyrhynchos No Canned beef luncheon meat, Bos taurus No Sliced ham, Sus scrofa Yes

6 6 Table S3: Analysis of specificity of DNA detection using B. subtilis primers with different bacterial samples Samples Detection of LAMP amplicons Normalized Relative intensity (%) Bacillus subtilis Yes Legionella pneumophila No Bacillus cereus No Salmonella enteric No Staphylococcus epidermidis No 100 Pseudomonas aeruginosa No Streptococcus pyogenes No

7 7 Table S4: Material and cost analysis of paper chip assay Material Cost (Cent) Cellulose paper 6 Wax 2 Primers 4.2 LAMP reagent 13.5 Chemical for surface chemistry 5 Total

8 8 Table S5 LAMP primers sets for amplification reaction. Cytochrome b gene in S. scrofa bp F3 5 - TCGCCTACGCTATTCTAC-3ʹ 18 B3 5 - GGAAGTATAAGATGGAGGCTA-3ʹ 21 FIP 5ʹ-GGATGTGTGTAGTATGGGCATTAACTAGGTGGAGTGTTGG-3ʹ 40 BIP 5ʹ-TTCGACCACTAAGTCAATGCCGGTTGTCCTCCAATTCATG-3ʹ 40 LF 5ʹ-ATTAGGATTAGGATGGAGGCTA-3ʹ 22 LB 5ʹ-CTAGTAGCAGACCTCATTACAC-3ʹ 22 rpob gene in B.subtilis F3 5ʹ-GAAGAGGATATGCCTTACCTTC-3ʹ 23 B3 5ʹ-CGACAGATACACGGTTATCAA-3ʹ 21 FIP 5ʹ-GGTAACGAGCGGCCATACCTACCATCACGTATGAACATCG-3ʹ 40 BIP 5ʹ-TGGCATTCACATTGCATCTCCTCCGGCTTCTTCAAGTGTT-3ʹ 40 LF 5ʹ-CATGTGAAGTTCCAATACCTGC-3ʹ 22 LB 5ʹ-GCGAGAAGAGGATGTCTGG-3ʹ 19 8

Salmonella in Food 2014

Salmonella in Food 2014 Salmonella in Food 04 Table : Salmonella spp. in meat Food Type Sampling Site RTE Fresh Meat Broiler Processing NS 0 0 Catering NS 0 0 Unspecified Poultry (Including Turkey, Duck and Unspecified NS 0 0