[timed_out] Timeout waiting for reply to request: LOAD_CONFIG

[kcleal]

Hi,

ReadBouncer keeps throwing this error whenever I start a sequencing run. Im using an old flow cell in live sequencing mode - it only has a few active pores, could this be a problem?

    IBF-build processed 2 sequences (0.000134 Mbp) in 0.00224295 seconds (53.5011 Kseq/m, 3.58457 Mbp/m)
     - 2 sequences in 2 bins were written to the IBF
    Trying to connect to MinKNOW
    Host : localhost
    Port : 9502
    Connecting to MinKNOW using the token: 7d2c7e22-bbfb-43b8-9d37-69fb6f90cf40
    Connection successfully established!
    Waiting for device to start sequencing!
    Please start the sequencing run now!
    Sequencing has begun. Starting live signal processing!
    Connecting to Guppy basecall server on address : 127.0.0.1:9502
    [guppy/error] basecall_service::BasecallClient::worker_loop: Connection error. [timed_out] Timeout waiting for reply to request: LOAD_CONFIG
    [timed_out] Timeout waiting for reply to request: LOAD_CONFIG
    Real time : 32.0663 sec
    CPU time  : 0.114984 sec
    Peak RSS  : 30 MByte

My config file is:

    [IBF]
    kmer_size          = 15         #(unsigned integer with default 13) only required for 'usage = "build"' or if target_file/deplete_file is a fasta formate file 
    fragment_size      = 100000     #(unsigned integer with default 100000)  only required for 'usage = "build" or if target_file/deplete_file is a fasta formate file 
    threads            = 3          #(unsigned integer with default 3)
    target_files  = ['/home/alex/Desktop/AdaptiveSampling/ref_seqs/16p1_21q.fasta']
    deplete_files = []
    #read_files    = ['path/to/read/file/SampleZMCDataSet.fasta']
    exp_seq_error_rate = 0.1        #(unsigned float between 0 and 1 default 0.1)
    chunk_length       = 360        #(unsigned integer with default 250)
    max_chunks         = 1          #(unsigned integer with default 5)

    [MinKNOW]
    host               = "localhost" #(ip address or name of the computer hosting MinKNOW)
    port               = "9502"      #(port number used fo grpc communication by by MinKNOW instance) 
    flowcell           = "MN42383"   #(name of the flowcell used)
    channels           = [1,512]

    [Basecaller]
    caller             = "Guppy" #DeepNano/Guppy (default is DeepNano)
    host               = "127.0.0.1"#(ip address or name of the computer hosting Guppy Basecall Server)
    port               = "9502"     #(port number on which the basecall server is running on the host)
    threads            = 3          # (unsigned integer with default 3)
    config             = "dna_r9.4.1_450bps_fast"

[JensUweUlrich]

I think you are using the wrong port for the basecaller here. On which host are you running the guppy_basecall_server? If you did not start a separate server instance, you should state the following.

[Basecaller]
caller = "Guppy"
host = "/tmp/.guppy"
port = "5555"
config = "dna_r9.4.1_450bps_fast"

[kcleal]

Hi, Thanks for the quick response. Im using my local machine - sequencing on the same machine as readbouncer. I will try this!

[kcleal]

It got passed that error, but I have a new zeromq error:

    Connecting to Guppy basecall server on address : /tmp/.guppy:5555
    [guppy/error] basecall_service::BasecallClient::worker_loop: Connection error. [failed] zmq::error_t : Invalid argument
    [failed] zmq::error_t : Invalid argument
    Real time : 311.907 sec
    CPU time  : 0.11749 sec
    Peak RSS  : 31 MByte

Do I need to change the Minknow port to 5555?

[JensUweUlrich]

Ah..sorry. That was my fault. Try this one:

[Basecaller]

caller             = "Guppy"  
host           = "ipc:///tmp/.guppy"
port               = "5555"
config         = "dna_r9.4.1_450bps_fast"

[kcleal]

Im still getting the same error unfortunately. The config file looks like:

  [Basecaller]
  caller             = "Guppy"
  host = "ipc:///tmp/.guppy"
  port = "5555"
  threads            = 3
  config             = "dna_r9.4.1_450bps_fast"

The output was:

  (base) alex@alex-ISRMOT-235:~/Desktop/AdaptiveSampling/experiments/Harsh_15.12.22/rbas$ ~/Desktop/AdaptiveSampling/software/ReadBouncer-1.2.2-Linux/bin/ReadBouncer --config ~/Desktop/AdaptiveSampling/experiments/Harsh_15.12.22/config.16p1_21q.toml
  IBF-build processed 2 sequences (0.000134 Mbp) in 0.0107952 seconds (11.116 Kseq/m, 0.744775 Mbp/m)
   - 2 sequences in 2 bins were written to the IBF
  Trying to connect to MinKNOW
  Host : localhost
  Port : 9502
  Connecting to MinKNOW using the token: f7ca0499-24e7-4172-9fd7-5a1e1943f995
  Connection successfully established!
  Waiting for device to start sequencing!
  Please start the sequencing run now!
  Sequencing has begun. Starting live signal processing!
  Connecting to Guppy basecall server on address : ipc:///tmp/.guppy/5555
  [guppy/error] basecall_service::BasecallClient::worker_loop: Connection error. [timed_out] Timeout waiting for reply to request: LOAD_CONFIG
  [timed_out] Timeout waiting for reply to request: LOAD_CONFIG
  Real time : 432.156 sec
  CPU time  : 0.1894 sec
  Peak RSS  : 31 MByte

Any more suggestions? Thanks

[JensUweUlrich]

Can you please tell me which Guppy version you are currently using on your machine?