SRX18286351 GSM6734822_r1 SRP408263 PRJNA902362 SRS15778123 GSM6734822 GSM6734822 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286352 GSM6734839_r1 SRP408263 PRJNA902362 SRS15778124 GSM6734839 GSM6734839 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286353 GSM6734840_r1 SRP408263 PRJNA902362 SRS15778125 GSM6734840 GSM6734840 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286354 GSM6734841_r1 SRP408263 PRJNA902362 SRS15778126 GSM6734841 GSM6734841 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286355 GSM6734842_r1 SRP408263 PRJNA902362 SRS15778127 GSM6734842 GSM6734842 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286356 GSM6734843_r1 SRP408263 PRJNA902362 SRS15778128 GSM6734843 GSM6734843 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286357 GSM6734823_r1 SRP408263 PRJNA902362 SRS15778129 GSM6734823 GSM6734823 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286358 GSM6734824_r1 SRP408263 PRJNA902362 SRS15778130 GSM6734824 GSM6734824 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286359 GSM6734825_r1 SRP408263 PRJNA902362 SRS15778131 GSM6734825 GSM6734825 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286360 GSM6734826_r1 SRP408263 PRJNA902362 SRS15778132 GSM6734826 GSM6734826 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286361 GSM6734827_r1 SRP408263 PRJNA902362 SRS15778133 GSM6734827 GSM6734827 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286362 GSM6734828_r1 SRP408263 PRJNA902362 SRS15778134 GSM6734828 GSM6734828 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286363 GSM6734829_r1 SRP408263 PRJNA902362 SRS15778135 GSM6734829 GSM6734829 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286364 GSM6734830_r1 SRP408263 PRJNA902362 SRS15778136 GSM6734830 GSM6734830 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286365 GSM6734831_r1 SRP408263 PRJNA902362 SRS15778137 GSM6734831 GSM6734831 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286366 GSM6734832_r1 SRP408263 PRJNA902362 SRS15778138 GSM6734832 GSM6734832 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286367 GSM6734833_r1 SRP408263 PRJNA902362 SRS15778139 GSM6734833 GSM6734833 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286368 GSM6734834_r1 SRP408263 PRJNA902362 SRS15778140 GSM6734834 GSM6734834 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286369 GSM6734835_r1 SRP408263 PRJNA902362 SRS15778141 GSM6734835 GSM6734835 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286370 GSM6734836_r1 SRP408263 PRJNA902362 SRS15778142 GSM6734836 GSM6734836 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286371 GSM6734837_r1 SRP408263 PRJNA902362 SRS15778143 GSM6734837 GSM6734837 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500 SRX18286372 GSM6734838_r1 SRP408263 PRJNA902362 SRS15778144 GSM6734838 GSM6734838 RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA Esophageal epithelium-enriched tissue layer was peeled from muscle then incubated in 1 ml 1X Dispase I (354235; Corning; Bedford, MA, USA) diluted 1:4 in Hank's Balanced Salt Solution (HBSS; 14025-076; Gibco; Grand Island, NY, USA) for 10 minutes at 37 °C with shaking at 1000 RPM (5384; Eppendorf F1.5 Thermomixer; Hamburg, Germany). Following removal from Dispase I, esophageal epithelium was minced with sharp scissors then incubated in 1 ml of 0.25% Trypsin-EDTA (25-053-CI; Corning; Manassas, VA, USA) for 10 min at 37 °C with shaking at 1000 RPM. Trypsin and tissue pieces were forced through 70 μm cell strainer into 50 ml conical tube containing 4 ml soybean trypsin inhibitor (STI; 9035-81-8; Grand Island, NY, USA). Cells were pelleted at 1300 RPM for 3 min then resuspended in 500 μl complete mouse keratinocyte–serum-free medium (37010022; Gibco; Carlsbad, CA, USA). Cell number and viability were measured using Automated Cell Count (AMQAX1000, Invitrogen Countess II, Bothell, WA, USA) by mixing 5 μl cell suspension with 5 μl 0.4% trypan blue solution (T10282; Invitrogen; Eugene, OR, USA). Library was performed according to the manufacter's instructions (Illumina inDrop). In brief, single epithelial cells were isolated into droplets and lysed. Droplets were fused with hydrogel microsphere containing cell-specific barcodes and underwent reverse transcription, with barcodes serving as primers. cDNA was purified from droplets and ligated to adapters, where they were amplified, annealed to indexed primers, and further amplified prior to sequencing. NextSeq 500