Kun:LabNotes/GenomeSeq/2013-6-15

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Data analysis of the third PGP1 cells[edit]

  • Hosuk did the amplification and amplicon extraction. He used a slower mixing and lower KOH concentration to avoid excessive DNA damage.
  • Noi constructed the libraries.
  • Sequencing was performed in HL154, 50% of Lane 4 and 100% of Lane 7, SE36+7 reads.
Library Raw_reads Mapping_rate Mappable_reads Clonal_rate 1+ 1 2 3 4+
HL154-HSL-PGP1-June4_Indx09 3,046,875 98.44% 2,999,294 0.8577 5,055,775 3,250,676 606,810 250,611 935,050
HL154-HSL-PGP1-June4_Indx10 2,839,916 95.24% 2,704,758 0.8424 4,066,194 2,600,783 421,754 169,135 857,959
HL154-HSL-PGP1-June4_Indx11 3,160,303 98.96% 3,127,378 0.9025 3,112,645 2,088,295 323,568 129,161 555,136
HL154-HSL-PGP1-June4_Indx12 2,650,212 98.57% 2,612,346 0.9134 2,534,151 1,765,505 280,636 105,155 373,716
HL154-HSL-PGP1-June4_Indx13 3,018,682 98.61% 2,976,825 0.908 2,828,285 1,916,281 284,036 118,674 497,474
HL154-HSL-PGP1-June4_Indx14 3,320,365 98.78% 3,279,773 0.9016 3,303,635 2,263,756 343,186 132,414 547,775
HL154-HSL-PGP1-June4_Indx15 1,777,529 98.22% 1,745,834 0.6706 5,842,769 3,578,417 729,552 288,223 1,230,259
HL154-HSL-PGP1-June4_Indx16 2,905,699 98.74% 2,869,141 0.6874 7,567,818 4,610,238 757,727 301,298 1,882,006
HL154-HSL-PGP1-June4_Indx17 2,569,434 98.42% 2,528,836 0.7446 5,323,875 3,241,562 501,135 208,936 1,355,721
HL154-HSL-PGP1-June4_Indx18 2,603,016 98.12% 2,554,130 0.6966 6,907,672 3,992,513 769,671 343,762 1,785,178
HL154-HSL-PGP1-June4_Indx19 2,508,037 98.21% 2,463,034 0.7099 7,050,967 4,311,747 824,038 332,709 1,565,919
HL154-HSL-PGP1-June4_Indx20 2,036,668 97.86% 1,993,023 0.6545 5,955,475 3,418,117 602,918 269,067 1,648,823
HL154-HSL-PGP1-June4_Indx21 2,687,448 98.64% 2,650,790 0.6586 7,532,080 4,416,498 746,925 321,270 2,030,841
HL154-HSL-PGP1-June4_Indx22 2,844,075 98.47% 2,800,692 0.6698 8,652,159 5,314,014 940,876 380,946 1,999,763
HL154-HSL-PGP1-June4_Indx23 3,161,089 96.92% 3,063,663 0.6949 8,040,240 4,449,145 867,736 401,182 2,305,614
HL154-HSL-PGP1-June4_Indx24 2,883,798 98.34% 2,836,002 0.7137 7,059,052 4,142,979 713,605 313,832 1,873,118
HL154-HSL-PGP1-June4_Indx25 1,906,394 97.70% 1,862,477 0.6118 6,560,072 3,759,050 675,301 314,092 1,795,346
HL154-HSL-PGP1-June4_Indx26 1,874,894 98.58% 1,848,223 0.6762 5,590,507 3,391,894 576,177 251,583 1,354,308
HL154-HSL-PGP1-June4_Indx27 1,906,657 98.80% 1,883,855 0.6614 5,381,553 3,186,537 522,240 229,564 1,426,705
HL154-HSL-PGP1-June4_Indx28 1,801,285 98.24% 1,769,633 0.6468 5,229,116 2,977,559 511,038 244,178 1,479,782
HL154-HSL-PGP1-June4_Indx29 2,265,674 97.27% 2,203,766 0.6576 6,666,084 3,859,133 684,057 303,194 1,803,181
HL154-HSL-PGP1-June4_Indx30 2,129,636 97.90% 2,084,867 0.6877 6,038,899 3,609,352 609,496 266,324 1,537,167
HL154-HSL-PGP1-June4_Indx31 3,148,226 98.63% 3,105,124 0.8398 6,722,352 4,346,394 897,073 364,993 1,097,451
HL154-HSL-PGP1-June4_Indx32 2,817,025 97.72% 2,752,791 0.8499 4,749,410 2,941,390 533,315 245,338 1,016,043
  File:HL154-HSL-PGP1-chr19.png
  • There are several issue with this batch of data:
    • First, based on the SeqMonk coverage plot, DNA amplification is extremely sparse, and the cross-talk among different reactors is stronger. The latter could be also due to the read mapping.
    • Second, there isn't clear evidence of big DNA molecules separated into individual reactors.
    • Third, the clonal rate is much higher, which is probably also due to the poor amplification.
    • Overall, there is something wrong with this experiment, either due to slow/incomplete mixing, or low KOH concentration. The next experiment needs to be design carefully to distinguish these two.