1：Experimental Design and Method Selection:

The study involved synthesizing four water-soluble cationic porphyrin derivatives with large side arm substituents, including an unreported octa-cationic porphyrin, and evaluating their ability to act as RLS probes for G-quadruplex recognition. Methods included UV-vis absorption, fluorescence, and RLS spectroscopy, circular dichroism (CD) spectroscopy, and linear discriminant analysis (LDA).

2：Sample Selection and Data Sources:

DNA oligonucleotides (e.g., ssDNA, dsDNA, G-quadruplexes like KRAS, Oxy28, Hum24, C-MYC, CatG4, AS1411) and microRNAs (e.g., Let-7a) were purchased from Sangon Biotech Co. Ltd. Concentrations were determined using absorbance at 260 nm with molar extinction coefficients.

3：List of Experimental Equipment and Materials:

Equipment included SHIMADZU RF-5301PC spectrofluorimeter for RLS and fluorescence spectra, Cary 60 UV-vis spectrophotometer (Agilent Technologies) for absorption spectra, BioLogic MOS-45 spectropolarimeter for CD spectra, Mercury Vx-500 spectrometer for NMR, Varian 7.0T FT-MS mass spectrometer for FT-MS, and Bruker Autoflex III TOF/TOF 200 instrument for MALDI-TOF-MS. Materials included cationic porphyrin derivatives, DNA oligonucleotides, Tris-HCl buffer, KCl, Na2EDTA, and various chemical reagents.

4：0T FT-MS mass spectrometer for FT-MS, and Bruker Autoflex III TOF/TOF 200 instrument for MALDI-TOF-MS. Materials included cationic porphyrin derivatives, DNA oligonucleotides, Tris-HCl buffer, KCl, Na2EDTA, and various chemical reagents. Experimental Procedures and Operational Workflow:

4. Experimental Procedures and Operational Workflow: DNA solutions were prepared in buffer, heated, cooled, and incubated. Porphyrin was added, and spectra were recorded. For RLS, excitation and emission monochromators were scanned synchronously at Δλ = 0 nm. Titration experiments varied DNA concentration with constant porphyrin concentration. Job plot analysis varied molar fractions. MicroRNA sensing involved mixing oligonucleotides, adding target microRNA, incubating, and recording RLS spectra.

5：Data Analysis Methods:

RLS intensity ratios (e.g., I702/I652) were used for ratiometric recognition. LDA was applied using combined data from UV-vis, fluorescence, and RLS spectroscopy with Matlab programming. Statistical methods included calculating detection limits based on linear relationships.