Automated priming and library loading device

Abstract

Provided herein are automated apparatus for the identification of microorganisms in various samples. The disclosure solves existing challenges encountered in identifying and distinguishing various types of microorganisms, including viruses and bacteria in a timely, efficient, and automated manner by sequencing.

What is claimed is: 
     
         1 . A method of reusing a flow cell in a sequencing reaction, the method comprising:
 (a) adding a first barcode to a first plurality of nucleic acid sequences from a first sample, thereby providing a first plurality of barcoded nucleic acid sequences;   (b) performing a first sequencing reaction on said first plurality of barcoded nucleic acid sequences on a sequencing apparatus comprising a flow cell;   (c) adding a second barcode to a second plurality of nucleic acid sequences from a second sample, thereby providing a second plurality of barcoded nucleic acid sequences; and   (d) performing a second sequencing reaction on said second plurality of barcoded nucleic acid sequences on said sequencing apparatus comprising said flow cell, thereby reusing said flow cell of said sequencing apparatus.   
     
     
         2 . The method of  claim 1 , wherein said first barcode and said second barcode are between 9 nucleotides and 18 nucleotides in length. 
     
     
         3 . The method of  claim 1 , wherein said first barcode and said second barcode are 18 nucleotides in length. 
     
     
         4 . The method of  claim 1 , wherein said first barcode and said second barcode are 9 nucleotides in length. 
     
     
         5 . The method of  claim 1 , wherein said first barcode and said second barcode have identical sequences. 
     
     
         6 . The method of  claim 5 , wherein said identical sequences form a periodic block design. 
     
     
         7 . The method of  claim 1 , wherein said first barcode and said second barcode have distinct sequences. 
     
     
         8 . The method of  claim 7 , wherein said distinct sequences form a nonperiodic block design. 
     
     
         9 . The method of  claim 1 , further comprising adding a third barcode to a third plurality of nucleic acid sequences from a third sample, thereby providing a third plurality of barcoded nucleic acid sequences. 
     
     
         10 . The method of  claim 9 , wherein said third plurality of nucleic acid sequences comprises a mixture of cDNA, RNA, and gDNA sequences. 
     
     
         11 . The method of  claim 9 , further comprising performing a third sequencing reaction on said third plurality of barcoded nucleic acid sequences, wherein said third sequencing reaction is performed on said sequencing apparatus comprising said flow cell, thereby repeating said reusing of said flow cell. 
     
     
         12 . The method of  claim 9 , wherein said first barcode, said second barcode, and said third barcode have identical sequences. 
     
     
         13 . The method of  claim 9 , wherein said first barcode, said second barcode, and said third barcode have distinct sequences. 
     
     
         14 . The method of  claim 11 , wherein said first barcode, said second barcode, and said third barcode have identical sequences. 
     
     
         15 . The method of  claim 11 , wherein said first barcode, said second barcode, and said third barcode have distinct sequences. 
     
     
         16 . The method of  claim 1 , further comprising: (e) performing an amplification reaction or nucleic acid enrichment on said first plurality of barcoded nucleic acid sequences prior to performing in (b). 
     
     
         17 . The method of  claim 1 , further comprising: (f) performing an amplification reaction or a nucleic acid enrichment on said second plurality of barcoded nucleic acid sequence prior to performing in (d). 
     
     
         18 . The method of  claim 1 , further comprising:
 (e) performing a first amplification reaction or a first nucleic acid enrichment on said first plurality of barcoded nucleic acid sequences prior to performing in (b); and   (f) performing a second amplification reaction or a second nucleic acid enrichment on said second plurality of barcoded nucleic acid sequence prior to performing in (d).   
     
     
         19 . The method of  claim 1 , wherein said first sequencing reaction or said second sequencing reaction distinguishes an epigenetic pattern on a nucleic acid from said first sample or said second sample, respectively. 
     
     
         20 . The method of  claim 19 , wherein said epigenetic pattern is a methylation pattern. 
     
     
         21 . The method of  claim 1 , wherein said first plurality of nucleic acid sequences or said second plurality of nucleic acid sequences comprises complementary DNA (cDNA) sequences. 
     
     
         22 . The method of  claim 1 , wherein said first plurality of nucleic acid sequences or said second plurality of nucleic acid sequences comprises ribonucleic acid (RNA) sequences. 
     
     
         23 . The method of  claim 1 , wherein said first sample, said second sample, or a combination of said first sample and said second sample is an environmental sample. 
     
     
         24 . The method of  claim 23 , wherein said environmental sample is obtained from a surface swab or a surface rinse of an environment, or wherein said environmental sample is obtained from a container, a piece of equipment, or a piece of clothing from a worker of an environment. 
     
     
         25 . The method of  claim 1 , wherein said sample is a biological sample that is not obtained from food, or wherein said biological sample comprises blood, plasma, urine, tissue, feces, bone marrow, saliva or cerebrospinal fluid of a subject. 
     
     
         26 . The method of  claim 1 , wherein said first plurality of nucleic acid sequences comprises a mixture of cDNA, RNA, and gDNA sequences, or wherein said second plurality of nucleic acid sequences comprises a mixture of cDNA, RNA, and gDNA sequences. 
     
     
         27 . The method of  claim 1 , further comprising demultiplexing, by a computer system, said second plurality of barcoded nucleic acid sequences comprising said second barcode from said first plurality of barcoded nucleic acid sequences comprising said first barcode. 
     
     
         28 . The method of  claim 27 , wherein said first sequencing reaction or said second sequencing reaction comprises a pore sequencing reaction. 
     
     
         29 . The method of  claim 1 , further comprising
 adding a third barcode to a third plurality of nucleic acid sequences from said first sample, thereby providing a third plurality of barcoded nucleic acid sequences; and   performing a third sequencing reaction on said third plurality of nucleic acid sequences.   
     
     
         30 . The method of  claim 29 , further comprising demultiplexing, by a computer system, said third plurality of nucleic acid sequences comprising said third barcode, said second plurality of nucleic acid sequences comprising said second barcode, and said first plurality of nucleic acid sequences comprising said first barcode.